R ES E A RC H
integration of these with systems biology tech-
REVIEW SUMMARY
BIOENGINEERING
Industrial biomanufacturing: The
future of chemical production
James M. Clomburg, Anna M. Crumbley, Ramon Gonzalez*
BACKGROUND: Environmental, geopolitical,
and economic factors are reshaping our view
of global energy and manufacturing demands.
Addressing these challenges may require a shift
from the current model of industrial chemical
manufacturing, which employs large-scale mega-
facilities that benefit from economies of unit
scale. Contrary to the traditional approach, a
model based on economies of unit number is
proposed to reduce capital costs per unit ca-
pacity. Industrial biomanufacturing, which ex-
ploits biological processes for manufacturing,
offers one way to address these changing global
factors while using the economies of unit num-
ber model. This model employs both facility-
level mass production of small-scale, modular
units and improvements to process design re-
sulting from repetition in a learning-by-doing
approach. The lower investment and financial
risk associated with smaller-scale, lowercapital
cost facilities allow a larger number and more
diverse group of technology players to be in-
volved, in turn enabling faster innovation and
novel technology adoption as well as a faster
response to market drivers.
ADVANCES: In contrast to current chemical
manufacturing methods, characteristics in-
herent to bioconversion processessuch as
the ability to operate at mild temperatures
and pressures and achieve high carbon- and
energy-conversion efficiencies in single-unit
operationsresult in more streamlined and
Methane-based industrial biomanufacturing for fuel and chemical production. Exploiting biological processes can enable the conversion of
single-carbon feedstocks, like methane, to the array of chemical products currently produced through industrial chemical manufacturing with con-
siderable economic, environmental, and societal advantages.
Clomburg et al., Science 355, 38 (2017) 6 January 2017
less technologically complex processes. These
characteristics enable flexible, smaller-scale, and
capital expenditureefficient operation that can
both support and benefit from a large number
of facilities, according to the economies of unit
number model. For example, the capital ex-
penditure entry-level cost of corn-grain ethanol
facilities, the most widely developed current
example of a bioconversion process, has sub-
stantially decreased as the number of plants
has increased over the past few decades. This
has facilitated rapid, small-scale, and wide-
spread deployment resulting in a more than
10-fold increase in U.S. ethanol production
from 1995 to 2015.
Advances in metabolic engineering, syn-
thetic biology, genomics, and industrial process
design have pushed industrial biomanufactur-
ing closer to more widespread adoption. Partic-
ular emphasis on single-carbon feedstocks,
such as methane or CO2, in applications where
large-scale chemical manufacturing is infea-
sible or too costly can leverage both econo-
mies of unit number for mass production of
facilities and the benefits of manufacturing
automation to reduce capital expenditure per
unit scale. Current research efforts focused on
the design of carbon- and energy-efficient meta-
bolic pathways have been particularly beneficial.
Advances in tool development for metabolic
pathway design have included in silico organism
design strategies, genome mining techniques,
and computational enzyme design efforts. The
niques, such as next-generation sequencing
and high-sensitivity omics methods, has al-
lowed for the design and potential development
of millions of chemical production pathways.
The use of genome engineering technologies
such as clustered regulatory interspaced short
palindromic repeats (CRISPR)associated pro-
tein Cas9 systems and multiplex automated
genomic engineeringand recent advances
in screening and select-
ON OUR WEBSITE ing for edited organisms
Read the full article using biosensors have re-
at http://dx.doi. duced the time required
org/10.1126/ to complete genomic edit-
science.aag0804 ing to a fraction of the
..................................................
traditional time require-
ments. Automation-based process design im-
Downloaded from http://science.sciencemag.org/ on February 16, 2017
provements of these biotechnology advances
have also facilitated rapid advances in indus-
trial biomanufacturing.
OUTLOOK: Continued development of indus-
trial biomanufacturing in the 21st century will
require further advances in biocatalyst design
and process design engineering. To facilitate a
future based on industrial biomanufacturing,
further development of genomic tools and in-
dustrial design automations suited to these
purposes is paramount. Furthermore, reduc-
ing the time required from concept design to
industrial relevance is essential. Specific de-
velopments in the area of one-carbon feed-
stocks stand to exploit the opportunity presented
by currently wasted, distributed methane through
the increased adoption of an economy of unit
numbers approach in industrial biomanufac-
turing. Although much work remains, the fu-
ture of industrial biomanufacturing holds great
promise in meeting the evolving demands of
chemical production in the current century and
beyond.
The list of author affiliations is available in the full article online.
*Corresponding author. Email: ramon.gonzalez@rice.edu
Cite this article as J. M. Clomburg, A. M. Crumbley, R. Gonzalez,
Science 355, aag0804 (2017). DOI: 10.1126/science.aag0804
1 of 1
R ES E A RC H

efficient and economically viable transportation

REVIEW infrastructure (Fig. 1B). For example, the 135
operating U.S. refineries produced more than
19.1 million barrels of oil per calendar day in 2015
BIOENGINEERING (8), with the majority of facilities operating with
a production capacity greater than 75,000 barrels

Industrial biomanufacturing: The per day (Fig. 1C). These large-scale facilities,
spanning upwards of 1375 ha (3500 acres) (Fig. 1A),
are located close to high-volume feedstock sources
future of chemical production (e.g., tight oil/shale gas) or transportation hubs
(e.g., Gulf coast) (Fig. 1B), ensuring high-volume
feedstock availability.
James M. Clomburg,1 Anna M. Crumbley,1 Ramon Gonzalez1,2*
The traditional model of economies of unit
scale described above for industrial chemical
The current model for industrial chemical manufacturing employs large-scale
manufacturing has long held that the only way
megafacilities that benefit from economies of unit scale. However, this strategy faces
to reduce capital costs per unit is by scale
environmental, geographical, political, and economic challenges associated with energy
essentially, bigger is better. Although this model
and manufacturing demands. We review how exploiting biological processes for
has been extremely successful, the inherent high
manufacturing (i.e., industrial biomanufacturing) addresses these concerns while also

Downloaded from http://science.sciencemag.org/ on February 16, 2017

capital expenditure, large scale, and long con-
supporting and benefiting from economies of unit number. Key to this approach is the
struction time result in a high financial risk that
inherent small scale and capital efficiency of bioprocesses and the ability of engineered
very few companies can tolerate, reducing the
biocatalysts to produce designer products at high carbon and energy efficiency with
number of new players able to gain entry. This
adjustable output, at high selectivity, and under mild process conditions. The biological
high financial risk accordingly hinders potential
conversion of single-carbon compounds represents a test bed to establish this paradigm,
transformative innovation by limiting the indus-
enabling rapid, mobile, and widespread deployment, access to remote and distributed
trial diversity and slowing the ability of the in-
resources, and adaptation to new and changing markets.
dustry to adapt to change. These characteristics

C
hemical manufacturing firms currently pro-
duce more than 70,000 products as part of
a worldwide industry projected to grow
beyond US$5 trillion by 2020 (1, 2). The
roots of chemical manufacturing are inher-
ently tied to the Industrial Revolution, a time
period in which a predominantly agrarian society
was transformed into one reliant on manufactur-
ing for domestic and worldwide markets. The
development and use of chemical processes was
a major part of this shift, as chemical techniques
for the large-scale production of compounds such
as sulfuric acid, bleaching powder, and soda ash
in the 18th century helped transform and shape
the production of glass, textiles, soaps, and paper
(35). These chemical processes laid the ground-
work for continued expansion in the 19th and
early 20th centuries, which would see the devel-
opment of manufactured fertilizers and petro-
chemicals that continue to be central to industrial
chemical manufacturing.
Advancing knowledge and understanding in
the field of biology in the 20th century enabled
the use of biological processes for the production
of important industrial compounds, such as ace-
tone and ethanol, in addition to many important
compounds in medicine, including penicillin and
other antibiotics. However, it was not until the
1970s, with the development of recombinant DNA
technologies, that the true potential of biological
processes for the industrial production of an array
of compounds was envisioned (6). From there,
developments in discovery, use, optimization, and
engineering of biological techniques, organisms,
and processes have led to a quickly evolving field
1
Department of Chemical and Biomolecular Engineering, Rice
University, Houston, TX, USA. 2Department of Bioengineering,
Rice University, Houston, TX, USA.
*Corresponding author. Email: ramon.gonzalez@rice.edu
for the commercial-scale production of both med-
ical and industrial compounds (7). Despite these
advancements, the earlier adoption, better un-
derstanding, and suitability of chemical tech-
niques for large-scale manufacturing of industrial
chemicals have made this industry essential for
our current way of life. However, the recent spot-
light on sustainability challenges inherent to in-
dustrial chemical manufacturing, coupled with
rapid advances in biotechnology, have brought
a renewed interest in industrial biomanufac-
turing. Here, we review recent advances related
to harnessing the power of biology for industrial
manufacturing, with a focus on applications pres-
enting major challenges for traditional chemical
techniques.
What are the limits to industrial
chemical manufacturing?
Current chemical manufacturing techniques,
which dominate fuel and chemical production,
employ processes incurring numerous heat and
pressure changes, with multiple unit operations
and heat integration schemes. Given that capital
infrastructure expenditures (CapEx) are largely
correlated to process complexity (i.e., amount
of energy and material transferred), the chem-
ical manufacturing industry has leveraged an
economies of unit scale model to reduce CapEx
per unit scale. This is reflected in both the large
CapEx and the immense scale and size of faci-
lities such as oil refineries, ethylene crackers,
ammonia plants, and gas-to-liquid (GTL) plants
(Fig. 1A). The economies of unit scale enabling
these megafacilities require large amounts of ag-
gregated feedstock(s)typically fossil-based, such
as petroleum and natural gasto support large-
scale production. These factors have led to a mod-
el in which a small number of large-scale facilities
rely on access to high-volume feedstocks and/or
may also restrain the suitability of the current
model to address the many interconnected eco-
nomic, environmental, geographical, and political
factors that are shifting the way we view global
energy and manufacturing demands in the 21st
century. For example, the requirement of direct
access to high-volume feedstocks inherently ties
industrial chemical manufacturing to large-scale
petroleum resources and limits feedstock diver-
sification. This not only limits access to fossil
resources such as remote natural gas but also
restricts the ability to shift to more sustainable
resources such as biomass, which are distributed
and smaller volume by nature (9). The large-scale
and energy-intensive nature of these facilities also
greatly contributes to their environmental impact
through wasted fossil resources during feedstock
extraction and chemical processing (10).
The cost, size, and scale characteristic of chem-
ical processing facilities mandated by the econo-
mies of unit scale model impose substantial
barriers to diversifying the players in chemical
manufacturing. This represents a concern for the
globalization of resources and the progression
of developing countries toward self-reliance for
chemical production. For example, the cost of
building a large-scale ammonia production plant,
oil refinery, or GTL facility (Fig. 1A) is on par with
the gross domestic product of countries like
Equatorial Guinea, Congo, and Gabon (US$8 to
15 billion) (11), countries that possess large oil
and natural gas reserves. This places an econo-
mic barrier on access to chemical manufacturing
facilities needed to use local resources for do-
mestic fertilizer or energy production, products
essential for progressive development. As such,
the characteristics of the economies of unit scale
model not only impose an access barrier to both
countries and companies but also limit the poten-
tial for rapid and innovative solutions to tackle
these challenges.

Clomburg et al., Science 355, eaag0804 (2017) 6 January 2017 1 of 10

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Fig. 1. Comparison of industrial

chemical manufacturing to indus-
trial biomanufacturing. (A) Capital
expenses (CapEx) versus capacity
for industrial chemical manufacturing
(oil refineries, steam crackers,
ammonia, and gas-to-liquids plants)
and industrial biomanufacturing
(corn-ethanol facilities and first-of-
their-kind 1,3-propanediol, cellulosic
ethanol, and 1,4-butanediol facilities)
on an equivalent energy basis
(BOE/day). Insets show satellite
images that illustrate differences in
footprint between a corn-ethanol
plant and an oil refinery. Inset values
highlight aerial productivity
(BOE/ha/hour), an aerial efficiency

Downloaded from http://science.sciencemag.org/ on February 16, 2017

calculation considering capacity and
facility footprint estimated from
Google Map Imagery. [Imagery
copyright 2016 Google; map data
copyright 2016 Google] (B)
Distribution of oil refineries (blue
flame) and corn-ethanol plants (green
leaf) in the United States. Production
capacity is in BOE/day. [Data from
references (90103); Mapping
and georeferencing copyright
OpenStreetMap and copyright Carto,
respectively] (C) Frequency
distribution of corn-ethanol plants
(green) and oil refineries (blue) in the
United States as a function of their
capacity on an equivalent energy
basis (thousand BOE/day).

Can industrial biological manufacturing
make the case for economies of
unit number?
Contrary to the traditional model of economies
of scale used in industrial chemical manufactur-
ing, a model based on economies of unit number
uses facility-level mass production and improve-
ments to process design resulting from repeti-
tion, a learning by doing approach, to reduce
capital costs per unit capacity (12). Here, eco-
nomies of unit number can be defined as a shift
from a small number of high-capacity units or
facilities to a large number of units or facilities
operating at a smaller scale. Through increasing
the number of units produced, this model can
leverage cost reduction through mass production
in which costs decline as cumulative output
increases because of specialization in the pro-
duction process and improved process and pro-
duct design (12). Furthermore, by increasing the
number of operating units and facilities, cumu-
lative experience on the process is gained that
can uncover a myriad of improvements in design,
materials, and production methods (12). These
continuous improvements can offer substantial
cost reductions as total production volume (i.e.,
number of facilities times facility production
rate) increases.
To exploit this economic model in the context
of chemical manufacturing, one can envision the
custom-built, high-CapEx, large-scale operations
being replaced with a large number of mass-
produced, modular, small-unit-scale technology.
This can enable an economies of unit number
approach that can use automation principles,
advances in engineering, and streamlining of
efficient technologies to reduce capital costs
per unit capacity. By efficiently scaling down,
this type of approach can support small-scale,
CapEx- and process-efficient flexible technolo-
gies. The smaller-scale and CapEx efficiency
reduces both the time and cost required for com-
mercial facilities, enabling rapid, mobile, and
widespread deployment. This model could address
many of the aforementioned limitations of in-
dustrial chemical manufacturing by enabling a
greater number of companies to access distributed
and sustainable resources and new markets and
allowing quick adaptation to both local and
global market conditions. Although historically,
available chemical technologies have not been
able to operate efficiently in such a fashion, the
development of technologies with lower overall
complexity that operate efficiently at a smaller
scale can help support mass-produced, modular,
small-unit-scale design.
In this context, exploiting the diversity and
strengths of biological processes for fuel and chem-
ical production provides the opportunity for CapEx
and process-efficient technology development,
enabling an economies of unit number model.
Industrial biomanufacturing employs biological
catalysts for the conversion of various feedstocks
to fuels and chemicals analogous to those cur-
rently produced via chemical manufacturing means
(Fig. 2). Biological processes, such as microbial
fermentation, efficiently operate at mild temper-
atures (20 to 100C) and pressures (atmospheric)
and can achieve high carbon- and energy-
conversion efficiencies in a single unit operation,
which in turn leads to more streamlined and less
technologically complex processes (13, 14). Indus-
trial biocatalysts afford the ability to produce a
single product with an easily adjustable output
at high selectivity without dramatically altering

Clomburg et al., Science 355, eaag0804 (2017) 6 January 2017 2 of 10

R ES E A RC H | R E V IE W
Fig. 2. Industrial biomanufacturing for fuel and chemical production. Exploiting biological processes can enable the conversion of numerous industrially
relevant feedstocks to the array of chemical products currently produced through industrial chemical manufacturing with considerable economic, environmental,
and societal advantages.
process conditions (15). As such, the process
equipment for the conversion step is largely
similar regardless of the desired output, which
can enable mass production of modular units
as opposed to custom-built equipment, both re-
ducing equipment cost and fabrication time.
These traits are ideally suited for a shift in chem-
ical production to a model based on economies
of unit number.
The potential for industrial biomanufacturing
to support economies of unit number can be
quantitatively established through the analysis
of corn-grain ethanol fermentation, which rep-
resents the most widely developed current ex-
ample of a bioconversion process adopted for
the production of chemicals and fuels at com-
mercial scale. Comparison of bioethanol plants
(industrial biomanufacturing) to oil refineries,
ethylene crackers, ammonia plants, and GTL
plants (industrial chemical manufacturing) on a
CapEx and barrels of oil equivalents (BOE) basis
demonstrates the extent to which industrial bio-
manufacturing can support smaller-scale, CapEx-
efficient facilities (Fig. 1A). This has resulted in
U.S.-based bioethanol production developing as
a network of distributed plants (Fig. 1B) that
operate on a substantially smaller scale than
chemical manufacturing facilities. The 195 opera-
tional bioethanol plants in the United States in
2016 have a total production capacity of 0.53 million
BOE/day, with the majority producing between
1000 and 5000 BOE/day (Fig. 1C). In contrast,
135 operating U.S. refineries produce more than
19.1 million barrels of oil per calendar day, about
35 times the total BOE produced by ethanol
plants, and operate with a production capacity
greater than 75,000 barrels per day (Fig. 1C),
which is 15 to 75 times the average production
capacity of ethanol plants. Despite differences
in production capacities, the reduced land usage
of bioethanol facilities enables aerial productiv-
Clomburg et al., Science 355, eaag0804 (2017) 6 January 2017
ities comparable to large-scale chemical oper-
ations (Fig. 1A). This system-level productivity
comparison further demonstrates how industrial
biomanufacturing facilities can effectively oper-
ate at a small scale, with process intensification
enabling further size reduction at comparable
magnitudes to that of chemical facilities.
The ability of these biomanufacturing facilities
for rapid, small-scale, and widespread deploy-
ment has supported a more than 10-fold increase
in U.S. ethanol production from 1995 to 2015
(16). Although one cannot overlook the role that
government incentives had in influencing the
ethanol market and demand, the intrinsic char-
acteristics (small-scale, CapEx efficiency) of these
bioconversion facilities enabled their rapid and
widespread deployment to capitalize on these
incentives and outpace the deployment of other
technologies. Furthermore, while corn-ethanol
represents the only current process with an es-
tablished network of facilities, other demonstrated
commercial-scale bioprocesses have CapEx per
unit capacities comparable to those of chemical
manufacturing facilities (Fig. 1A). DuPonts first-
of-their-kind bio-1,3-propanediol and cellulosic eth-
anol facilities operate at similar CapEx/capacity
metrics to that of many recently constructed re-
fineries and ethylene crackers, with the Novamont
1,4-butanediol plant, using Genomatica engineered
strains for bioconversion, displaying slightly higher
CapEx/capacity than would be expected of a first-
of-its-kind process.
Central to the rapid adoption of these bio-
manufacturing technologies is the fact that the
CapEx entry-level cost of ethanol production via
fermentation has markedly decreased over the
past few decades. Focusing on capital expend-
iture costs, which are representative of stan-
dard biological production processes, rather than
process-specific operational costs, the decreasing
CapEx cost has been the result not of increas-
Downloaded from http://science.sciencemag.org/ on February 16, 2017
ing the scale of facilities per se, but rather in-
creasing the number of facilities built (Fig. 3).
Although modest cost gains have been achieved
by optimizing the scale of the facilities, techno-
logical learning has had a dramatic effect on
CapEx reduction by both reducing costs related
to production and energy and driving process
optimization and integration. As a result of learn-
ing by doing, the CapEx per unit capacity of corn-
ethanol production facilities of the same scale
has dramatically decreased over the past 30 years,
affording industrial biomanufacturing processes
the ability to operate at a small scale in a CapEx-
efficient manner (Fig. 3). This analysis suggests
that not only do industrial biomanufacturing pro-
cesses require lower capital investment and oper-
ate in a CapEx-efficient manner at a smaller scale
(Fig. 1A) but also these processes support and
benefit from an abundance of small-scale facili-
ties according to the economies of unit number
model rather than the established paradigm of
economies of unit scale for industrial chemical
manufacturing (Fig. 3).
The lower investment and financial risk asso-
ciated with smaller-scale industrial biomanufac-
turing facilities allows a larger number and more
diverse group of technology players to be involved,
in turn enabling faster innovation and novel tech-
nology adoption as well as a faster response to
market drivers. The ability for rapid and distrib-
uted deployment enables feedstock diversifica-
tion by enabling access to remote, smaller-scale
resources. This can not only help reduce envi-
ronmental impact by capitalizing on sustain-
able feedstocks such as biomass but also help
reduce greenhouse gas emissions through tar-
geting smaller-scale methane resources like flared
and vented natural gas and biogas. The smaller-
scale, lower-CapEx facilities can also promote
equitable resource globalization by lowering
the economic barrier to chemical manufacturing.
3 of 10
R ES E A RC H | R E V IE W
Fig. 3. Reduction in capital costs of U.S. etha-
nol plants since 1977. Capital costs have been
reduced by upscaling, but even more by learning
and other nonscale-related development. Feasibil-
ity study markers have hollow center. [Adapted
from (104). Data obtained from (104111).]
For example, a lower economic entry threshold
removes a substantial burden on developing
countries desiring to drive economic develop-
ment, such as in the sub-Saharan Africa natural
gas reserve region, through capitalization on local
resources for domestic fertilizer and energy pro-
duction and minimization of import products.
As such, the use of industrial biomanufactur-
ing can enable the democratization of chemical
production.
How feasible are single-carbon
feedstocks as a test bed for
industrial biomanufacturing?
Although the aforementioned characteristics of
industrial biomanufacturing are being recognized,
widespread adoption may be facilitated by suc-
cessful implementation in a specific sector where
large-scale chemical manufacturing is not feasi-
ble. This could be due to small volumes, the dis-
tributed nature of the feedstock, inherently high
technological complexity, inefficiency of chemi-
cal processing, or general market isolation. The
production of organic chemicals from single-
carbon (C1) feedstocks, especially methane, offers
one such opportunity for industrial biomanu-
facturing to overcome these challenges. Natural
gas associated with petroleum extraction, as well
as methane from agricultural and landfill waste
decomposition and wastewater treatment, are
small-scale, highly distributed sources of meth-
ane (Fig. 4A), which do not represent a feasible
feedstock for large-scale chemical manufacturing
facilities. A quantitative analysis of the amounts
of methane generated from flared natural gas,
agricultural sites, and landfill waste demonstrates
the feedstock limitation challenge for current
large-scale chemical facilities and the oppor-
tunity for application of small-scale industrial
biomanufacturing technologies. On a BOE ba-
sis, these resources generate substantially lower
amounts than required for oil refinery feed-
Clomburg et al., Science 355, eaag0804 (2017) 6 January 2017
stocks; however, the amounts produced are on
par with ethanol plants on a BOE basis (compare
Fig. 1C and Fig. 4B). For example, gas-generation
rates from 1233 U.S. Environmental Protection
Agency (EPA)reporting landfill projects range
between 2 and 300 BOE/day (Fig. 4B), whereas
gas production from 239 EPA-reporting agricul-
tural sites operate at a narrower range centered
around 2 to 20 BOE/day (Fig. 4B). Flaring from
490 petroleum operations visible through the
National Oceanic and Atmospheric Administra-
tions Visible Infrared Imaging Radiometer Suite
(VIIRS) Nightfire (17) indicates the smaller scale
of flare-associated gas as well, with the majority
of sites generating 2 to 40 BOE/day (Fig. 4B).
Despite the small scale of individual-site meth-
ane generation, the large number of sites results
in substantial total resources, with more than
250 billion cubic feet/year generated from vent-
ing and flaring and nearly 240 billion cubic feet/
year from landfill, wastewater, and agricultural
associated sites (Fig. 4). This represents a major
opportunity for both U.S. and global fuel and
chemical production. In fact, the conversion of
all carbon associated with the more than 4 tril-
lion cubic feet of natural gas vented or flared
worldwide in 2014 is enough to meet global
production of seven essential building-block or-
ganic chemicals (methanol, ethylene, propylene,
butadiene, xylene, benzene, and toluene) that are
currently used for the production of hundreds of
chemicals (2). Despite this potential, the dis-
tributed and small-scale nature of methane re-
sources remains a major challenge for use via
current chemical technologies, as methane is
difficult to chemically process at mild condi-
tions and requires complex techniques. The eco-
nomics of natural gas recovery require a given
rate of methane production to justify the in-
frastructure expenditures necessary to gather,
process, and transport volumes of gaseous meth-
ane (Fig. 4C) (18). Although complex, large-scale
techniques are viable for high production rates
or short distances to market, the remote locations
of many oil and gas production sites strain tra-
ditional economic recovery justification, because
no current conversion technology can efficiently
access the lower-volume production sites located
at far distances from market hubs (Fig. 4C). This
results in a large amount of stranded (remote)
natural gas, a wasted resource due to the unfav-
orable economics of technically complex, large-
scale chemical processes.
Given that the traditional industrial chem-
ical manufacturing characteristics of large scale,
high CapEx, and long construction time are not
amenable to accessing these small-scale, distrib-
uted resources, this scenario represents an ideal
test case for the application of an economies of
unit number approach. Feedstock availability in
small amounts at a large number of sites will
require a large number of distributed, small-scale
facilities to capitalize on the available resource,
which is currently wasted due to technical and
economic incompatibility with existing methods.
This enables a learning-by-doing approach that
can leverage both economies of unit number for
mass production of facilities and the benefits of
manufacturing automation to reduce CapEx per
unit scale. Additionally, distributed feedstocks
can be coupled to localized markets by tailor-
ing product synthesis to regional market needs.
The aforementioned characteristics of indus-
trial biomanufacturing (e.g., small-scale, flexible,
CapEx-efficient processes) supports this approach
and can provide the needed process and econo-
mic characteristics to recover methane resources
through rapid, mobile, and widespread technol-
ogy deployment directly at the feedstock source.
Recovering these distributed gas feedstocks al-
lows capitalization on otherwise wasted or in-
accessible resources, which if left to current
technologies will continue to result in consider-
able greenhouse gas emissions.
Although the demonstrated abilities of bio-
Downloaded from http://science.sciencemag.org/ on February 16, 2017
conversion processes to operate in a small-scale,
CapEx-efficient manner places industrial bio-
manufacturing in a unique position to use these
resources, several important biocatalyst and pro-
cess considerations must first be addressed. On
the process side, despite the success of bioethanol
facilities, the process equipment (e.g., reactors),
separation methods and unit operation(s), and
waste-stream management needed for gas-intensive
fermentation present different requirements from
those established for commercial bioconversion
processes. However, the widespread nature of
remote methane resources that are difficult to
access using chemical techniques can enable
exploitation of the learning-by-doing approach
to address these process requirements. By ac-
cumulating process knowledge and improving
and optimizing technologies, CapEx costs can
be reduced as the total number of facilities and
production volume increases in a manner sim-
ilar to that demonstrated by bioethanol plants.
This not only will further advance bioconversion
technology specific to methane use but also can
provide continuous knowledge that can be lever-
aged for other bioconversion applications target-
ing different feedstocks or complex products.
Although current research on aspects such as
gas transfer, continuous process operation, and
product separation in the context of bioconver-
sion processes are facilitating initial process ad-
vancements, improvement in biocatalyst design
represents another critical factor. Developing
biocatalysts exhibiting high conversion rates and
efficiencies is a central component that must
be improved to enable the required small-scale,
CapEx-efficient processes to capitalize on the op-
portunity presented by remote methane resources.
How can metabolic engineering enable
the biomanufacturing of fuels and
chemicals from C1 feedstocks?
Regardless of whether chemical or biological
methods are used, overcoming the high stability
and low reactivity of an inert CH bond is a major
challenge for the conversion of methane. Biolog-
ically, the vast majority of organisms known to
use methane have accomplished this through
oxygen-dependent CH bond activation, cata-
lyzed by methane monooxygenases (MMOs), in
4 of 10
R ES E A RC H | R E V IE W

Fig. 4. Methane resources are

highly distributed, small-scale
feedstocks that are challenging
for use by large-scale chemical
facilities. (A) Generation of meth-
ane at natural gas (red flame),
agricultural biogas (light blue
grass), and landfill (yellow trash
bag) facilities in the United States.
Relative size of icon correlates with
production (in BOE) at each site.
[Data from references (90103);
mapping and georeferencing
copyright OpenStreetMap and
copyright Carto, respectively] (B) Gas
production rate and BOE equivalency
distribution from 1233 EIA-reporting
landfills with capture/flaring capabil-

Downloaded from http://science.sciencemag.org/ on February 16, 2017

ities (88,779.25 BOE/day), 239 EIA-
reporting agricultural sites with biogas
generation capabilities (1597.434
BOE/day), and 490 visible VIIRS-
identified petroleum flaring wells
(16,118.20 BOE/day). [Data from
references (90103)] (C) Economics
of natural gas transport. Indicated
technologies for a given area repre-
sent economically favorable technol-
ogies for transports. Although
technologies are available for high
production rates and/or short
distances to market, the combination
of low production rates and large
distances to markets results in an
economically infeasible situation
resulting in stranded gas. Modified
from (18). CNG, compressed natural
gas; LNG, liquefied natural gas; GTL,
gas-to-liquid; NGH, natural gas
hydrates.

which a CH bond in methane is converted to a
COH group. Two variations of MMO exist: par-
ticulate, membrane-bound MMO (pMMO), and
cytoplasmic, soluble MMO (sMMO) and, given
their role and novel function in methanotrophs,
these enzymes have been extensively studied
(19). Obligate methanotrophs metabolize meth-
ane as their sole carbon and cellular energy
[adenosine triphosphate (ATP)] source, oxidizing
methane to CO2 via the intermediates metha-
nol (CH3OH), formaldehyde (CH2O), and formate
(CHO2 ). Methane-derived carbon is assimilated
at the level of formaldehyde via the ribulose-
monophosphate (RuMP) cycle or serine cycle,
formate via the serine cycle, or CO2 via the Calvin-
Benson-Bassham (CBB), depending on the specific
metabolism (Fig. 5) (20, 21). The majority of
methanotrophs are organized into two groups,
designated Gammaproteobacteria or Group I, which
use the RuMP pathway for carbon assimilation,
and Alphaproteobacteria or Group II, which use
the serine cycle (22). More recently, thermoacido-
philic aerobic methanotrophs (Verrucomicrobia)
have been identified, which can assimilate car-
bon through the CBB cycle at the level of CO2 (23).
Efforts to identify methanotrophic organisms
for industrial fermentation have revealed sev-
eral promising directions, despite historical li-
mitations due to low organism productivity and
a lack of genetic tools for manipulation. Re-
cent efforts have focused on Methylomicrobium
buryatense, a Group I methanotroph with a faster
doubling time compared with traditional meth-
anotrophs (24, 25). Genetic tools have recently
been developed for M. buryatense (26, 27) and
the ability for transient gene expression opens
the door for the development of powerful gen-
etic tools such as clustered regulatory interspaced
short palindromic repeats (CRISPR)/CRISPR-
associated protein Cas9based genome editing.
Recently, application of rational metabolic engi-
neering strategies employing these genetic engi-
neering tools enabled the development of a strain
of M. buryatense engineered for the production of
lactate from methane. Heterologous overexpres-
sion of a Lactobacillus helveticus L-lactate de-
hydrogenase in M. buryatense enabled L-lactate
production from methane, with cultivation in a
continuous bioprocess resulting in the produc-
tion of 0.8 g/L L-lactate (28).
Methanotrophic bacteria have also been used
for the production of a number of compounds
from methane, including BioProtein (2931), poly-
hydroxybutyrate (32, 33), carotenoids (34), vita-
mins (35), and methanol (36) (Table 1). Recently,
the identification of an active Embden-Meyerhof-
Parnas (EMP) pathway in novel gammaproteo-
bacterial methanotrophs, notable for resistance
to the toxic components of natural gas and bio-
gas, has redefined carbon efficiency calculations
and ATP availability in certain methanotrophic
organisms, most notably indicating the opportu-
nity for organic acid production via fermentation
(37, 38). EMP pathway presence provides the po-
tential to couple methanotrophic use with con-
ventional industrial strain-engineering routes
from sugars to biochemical intermediates and
products due to the conserved downstream meta-
bolic machinery (Fig. 5). Continued pathway dis-
covery, advancements in genetic tools, and focused
metabolic engineering efforts in the methano-
trophic space stand well-placed to further ex-
pand the number of chemicals produced and
improve carbon conversion efficiency. Improv-
ing both the range of products and the efficiency

Clomburg et al., Science 355, eaag0804 (2017) 6 January 2017 5 of 10

R ES E A RC H | R E V IE W

of this activation mechanism is limited by the

basic electron requirement needed to reduce
O2 to H2O. Limitations of electron flow inherent
to currently available MMO enzymes require
that electrons used in the oxidation of methane
to methanol be recovered in subsequent steps,
limiting the overall energy conversion efficiency.
Although directly coupling electrons generated
from methanol oxidation to methane oxidation
provides the most efficient MMO-based route
(40), exploiting alternative, theoretically more
efficient, activation routes is one way to achieve
improved energy efficiency. Several alternative O2-
dependent activation routes have been proposed
that have the potential to reduce the electron re-
quirement for methane activation (13). However,
these strategies have yet to be demonstrated.
One alternative to maximize energy efficiency

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is through the use of an anaerobic methane
activation mechanism. Anaerobic oxidation of
methane (AOM) is carried out by anaerobic meth-
anotrophic (ANME) archaea capable of coupling
sulfate reduction to methane oxidation to drive
favorable thermodynamics. AOM is a poorly un-
derstood biological process driven by natural
ANME and syntrophic bacteria consortia. De-
spite decades of effort, neither a natural consor-
tia nor pure microbial culture that grows on
methane anaerobically has been successfully iso-
Fig. 5. Metabolic pathways for the conversion of C1 feedstocks to fuels and chemicals. Leveraging lated (13, 41, 42). However, strong evidence sug-
recent developments in synthetic and systems biology enable the exploitation of various activation and gests that the enzyme responsible for the first
utilization pathways along with engineering integrated product synthesis pathways for the conversion of step of AOM is a homolog of the methyl-coenzyme
C1 feedstock to desired products. Continued understanding and development of mechanisms and M reductase (MCR) from anaerobic methanogenic
organisms for C1 use can enable improved bioconversion efficiency. Formaldehyde is assimilated directly microorganisms (42, 43), and reaction intermed-
through the RuMP pathway (blue) and is channeled to cellular metabolism and product formation via the iates and a proposed mechanism have recently
EMP pathway (gray). Formaldehyde and formate can be assimilated via 5,10-methylene-tetrahydrafolate been identified (44). These discoveries can aid in
(5,10-MTHF) through the Serine cycle (red) and are linked to cellular function through generation of acetyl- the development of technologies for novel meth-
CoA and biosynthesis in the cell via the glyoxylate cycle (purple). The Calvin-Benson-Bassham cycle ane activation. This type of approach was re-
(green) is responsible for carbon fixation in photosynthetic organisms and is tied to cellular metabolism cently demonstrated through the cloning of the
through production of pyruvate (PYR) and acetyl-CoA. Additional pathways for activation of methane and MCR from an unculturable organism, anaerobic
methane-derived substrates include the MCR methanogenesis reversal pathway (orange) and the com- methanotrophic archaeal population 1 (ANME-1)
putationally designed FLS enzyme pathway (pink) for the conversion of formaldehyde to dihydroxyacetone from a Black Sea mat, into the archaeal meth-
(DHA). Rationally engineered products are boxed. Dihydroxyacetone phosphate, DHAP; phosphoenolpyr- anogen Methanosarcina acetivorans to effec-
uvate, PEP; ribulose-5-phosphate, Ru5P; ribulose-1,5-phosphate, Ru1,5P2; 3-phosphoglycerate, 3PG; 1,3- tively run methanogenesis in reverse (41). Upon
biphosphoglycerate, 1,3P2G; glyceraldehyde 3-phosphate, G3P; fructose 1,6-biphosphate, F1,6P2; fructose coupling to iron reduction, engineered M. ace-
6-phosphate, F6P; xylose 5-phosphate, X5P; ribose-5-phosphate, R5P; sedoheptulose 7-phosphate, tivorans was able to grow anaerobically on meth-
S7P; sedoheptulose 1,7-biphosphate, S1,7P2; erythrose 4-phosphate, E4P; 5,10-methylene tetrahy- ane as a pure culture and convert methane into
drofolate, 5,10-MTHF; serine, SER; hydroxypyruvate, HPYR; glycerate, GLYC; 2-phosphoglycerate, acetate. ANME cultures have also been shown
2PG; 3-phosphoglycerate, 3PG; oxaloacetate, OAA; malate, MAL; glyoxylate, GLYOX; glycine, GLY; citrate, to decouple from their syntrophic bacterial part-
CITR; cis-aconite, cisA; succinate, SUCC; acyl-acyl carrier protein, Acyl-ACP; free fatty acid, FFA; ners and perform methane oxidation using sol-
fatty acid methyl ester, FAME; farnesyl pyrophosphate, FPP; geranylgeranyl pyrophosphate, GGPP; uble artificial electron acceptors (45), which holds
hydroxymethylglutaryl-CoA, HMG-CoA; (R)-3-hydroxybutyryl-CoA, (R)-3-HB-CoA; poly(3-hydroxybutyrate), great promise for the use of pure cultures in meth-
PHB. Pathways obtained from (41, 112115). ane bioconversion. More recently, the pathway
responsible for the synthesis of coenzyme F430, a
nickel-containing tetrapyrrole required for MCR
activity, was reconstructed in E. coli, opening the
of conversion is central to the development of gineering. However, attempts to transfer methane door for metabolic engineering efforts based on the
biocatalysts able to support the small-scale, oxidation machinery into heterologous hosts have use of MCR (46). These recent advances in meth-
CapEx-efficient processes needed to exploit avail- proven difficult. Only the soluble domain of the ane activation and energy-efficient pathways for C1
able methane resources and further advance the beta subunit of sMMO has been expressed in E. conversion continue to expand the potential for
potential for industrial biomanufacturing. coli (39), and heterologous expression of a com- developing more efficient biocatalysts target-
The use of model industrial microorganisms, plete, functionally active, pMMO has not been ing the production of fuels and chemicals from
such as Escherichia coli and Saccharomyces reported to date. methane.
cerevisiae, for a methane bioconversion process Even as industrial application of organisms Although methane represents the potentially
is of great interest given their rapid growth rates, using MMO-based methane activation offers most transformative C1 case to further enable
modern molecular techniques, and extensive lib- an attainable route for the conversion of meth- industrial biomanufacturing to target distrib-
rary of knowledge enabling rapid metabolic en- ane to chemical products, the energy efficiency uted, small-scale resources, biological processes

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R ES E A RC H | R E V IE W

Table 1. Biological conversion of single-carbon feedstocks to fuels and chemicals.

C1 feedstock Chemical product Titer Organism Reference

Methane L-Lactate 0.8 g/L Methylomicrobium buryatense (28)

............................................................................................................................................................................................................................................................................................................................................
Methane Astaxanthin 2.4 mg/g cell dry weight Methylomonas sp. 16A (34)
............................................................................................................................................................................................................................................................................................................................................
Methane Polyhydroxybutyrate 0.6 g PHB/g cell dry weight Methylocystis parvus OBBP (116)
............................................................................................................................................................................................................................................................................................................................................
Methane Methanol 1.1 g/L Methylosinus trichosporium OB3b (117)
............................................................................................................................................................................................................................................................................................................................................
Methanol Vitamin B12 800 ng/g wet biomass Methylobacterium sp. G-10 (35)
............................................................................................................................................................................................................................................................................................................................................
Methanol (R)-3-hydroxybutyrate 2.8 g/L Methylobacterium rhodesianum (118)
............................................................................................................................................................................................................................................................................................................................................
Methanol L-glutamate 60 g/L Bacillus methanolicus MGA3 (119)
............................................................................................................................................................................................................................................................................................................................................
Methanol L-lysine 65 g/L Bacillus methanolicus M168-20 (120)
............................................................................................................................................................................................................................................................................................................................................
Methanol Cadaverine 11.3 g/L Bacillus methanolicus (121)
............................................................................................................................................................................................................................................................................................................................................
Methanol a-humulene 1.65 g/L Methylobacterium extorquens AM1 (122)
............................................................................................................................................................................................................................................................................................................................................
CO 2 + electricity Isobutanol and 3-methyl-1-butanol 0.14 g/L total Ralstonia eutropha H16 (52)
............................................................................................................................................................................................................................................................................................................................................
Syngas n-Butanol 0.148 g/L Clostridium ljungdahlii (58)
............................................................................................................................................................................................................................................................................................................................................

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CO 2 + H 2 Acetone 3.02 g/L Acetobacterium woodii (63)
............................................................................................................................................................................................................................................................................................................................................
CO + H 2 2 Butyrate 1.3 g/L Clostridium ljungdahlii (61)
............................................................................................................................................................................................................................................................................................................................................

can also be exploited to use other single-carbon processes. As the most oxidized C1 feedstock, CO2 of such organisms (5860). Further efforts have
feedstocks, including methanol, formaldehyde, represents a single-carbon substrate that requires enabled the conversion of syngas feedstocks to
formate, and carbon dioxide. C1 feedstocks not very different mechanisms for use. The high level an array of products, including potential fuels,
only hold great promise for a number of appli- of oxidation requires energy input to produce more biopolymers and precursors, specialty plastics,
cations (31) but also demonstrate the flexibility of reduced carbon forms, and as a result the biological nylon precursors, solvents, and pharmaceutical
biocatalysts and bioconversion processes to tar- fixation of CO2 proceeds through a number of precursors, among others (5964). Benefits of
get various feedstocks. As an intermediate of different enzymes independent of those discussed bioconversion syngas fermentation include bio-
aerobic methane activation, the use of methanol above. Pathways for CO2 fixation include the catalyst tolerance to gas impurities and flexibility
as a carbon source requires similar pathways to Calvin-Benson-Bassham cycle, the reductive tri- regarding gas feedstock composition, whereas
that of methane, and many microorganisms carbocylic acid cycle, the Wood-Ljungdahl pathway, challenges include gas-liquid mass transfer li-
capable of using methane are also able to use the 3-hydroxypropionate:4-hydroxybutyrate cycle, mitations and the availability of genetic tools
methanol. Accordingly, organisms have been and the dicarboxylate:4-hydroxybutyrate cycle (55, 56). Despite these challenges, the potential
engineered to produce compounds from meth- (50). Each uses different key CO2 fixation enzymes, for syngas fermentation through combinatorial
anol (Table 1). Furthermore, identification and and these pathways are found across a wide scope metabolic engineering and process development
characterization of enzymes involved in meth- of various microorganisms. Microalgal species and has recently been demonstrated by the large-
anol use have enabled the construction of path- cyanobacteria are candidates for the conversion of scale production of ethanol (64), also demon-
ways for methanol conversion both in vitro (47) CO2 to various products, and the availability of strating that the challenges of gas-intensive
and heterologously in vivo (48). Notably, a unique genetic tools has enabled the metabolic engineer- fermentations can be overcome.
approach for the fixation of general C1 units ing of cyanobacteria, including Synechococcus Unique approaches for providing the required
has recently demonstrated a computationally elongatus PCC 7942 and Synechocystis sp. PCC reducing power for CO2 use have also been ex-
designed enzyme, formolase (FLS), which cat- 6803, to produce a number of compounds di- plored. One such approach is the use of microbial
alyzes the carboligation of three one-carbon rectly from CO2 (51). Despite this success, a major biofilms that directly accept electrons from elec-
formaldehyde molecules into one three-carbon challenge with the use of photosynthetic or- trodes for the reduction of CO2 to organic products
dihydroxyacetone molecule (49). This approach ganisms is providing large light-exposing sur- (65), a process referred to as microbial electro-
enabled the conversion of formate to the central faces, which requires extensive process design synthesis. Clostridium ljungdahlii has been shown
carbon metabolite dihydroxyacetone phosphate and optimization. to be highly effective in electrosynthesis for
in vitro, providing a proof-of-principle demon- An alternative to light-energy, fixation of the acetate production (65), with a newly developed
stration for the potential of a FLS pathway to oxidized carbon in CO2 can also be accom- genetic toolbox making it feasible to expand
convert C1 feedstocks such as formate or for- plished through the input of reducing power in product synthesis from CO2 (66). Another in-
maldehyde into chemical products. In addition the form of electrons from shuttles such as H2 novative method for direct electron input to
to providing routes to product synthesis from and formic acid. Systems in which H2, either the Wood-Ljungdahl pathway of an acetogen
these C1 feedstocks, these reports also represent externally added or produced via electrocata- has recently been reported in which the microbes
important steps in transferring methanotrophic lytic water splitting, or electrochemically pro- are photosensitized (67). Self-photosensitization
ability because these pathways can be combined duced formate, from water and CO2, are coupled of a nonphotosynthetic bacterium, Moorella
with continued efforts for heterologous MMO to the growth of lithoautotrophic organisms, thermoacetica, was demonstrated in which bio-
expression to enable various routes to metabolic have been demonstrated as a means of convert- logically generated cadmium sulfide semicon-
intermediates and product synthesis from methane. ing CO2 to useful chemical products (5254). Sim- ducting nanoparticles absorb light and use the
Developing and improving pathways for the ilarly, anaerobic fermentation of syngas, a fuel energy to carry out photosynthesis for the pro-
use of the above C1 compounds can aid in the gas mixture of CO2, H2, and CO, by acetogenic duction of acetate from CO2.
development of efficient biocatalysts for tar- bacteria via the Wood-Ljungdahl pathway, has Given the challenge of reducing the highly
geting distributed methane feedstocks. In addition, been demonstrated for the production of fuels oxidized carbon in CO2, continued insight into
given the gaseous nature of CO2, bioconversion and chemicals from C1 feedstocks (5557). A num- the characterization and mechanism of enzymes
processes targeting this feedstock offer the po- ber of acetogens have been used for this pur- for CO2 reduction is another critical area of re-
tential to both contribute to and leverage from pose, with the recent development of genetic tools search. Despite being a widely distributed reaction
knowledge gains for gas-intensive fermentation a key step in enabling the metabolic engineering in nature, relatively little is known about enzymes

Clomburg et al., Science 355, eaag0804 (2017) 6 January 2017 7 of 10

R ES E A RC H | R E V IE W
catalyzing CO2 reduction activity. Recently, a
novel, hydrogen-dependent CO2 reductase, pur-
ified from the model acetogenic organism Ace-
tobacterium woodii, was identified that fixes CO2
to formate via the electron shuttle H2 (68, 69).
Furthermore, the ability for enzymes such as
formate dehydrogenase to catalyze the reduc-
tion of CO2 has also been discovered (70), and
further characterization and discovery can be ex-
ploited to better understand and improve CO2
utilization processes. Of special interest is the
form of the electron donor, which can potentially
be coupled to other biological reactions to max-
imize overall efficiency.
What is the future of
industrial biomanufacturing?
Despite the promise of C1 feedstocks for indus-
trial biomanufacturing of fuels and chemicals,
much work remains to develop biocatalysts ex-
hibiting the required overall carbon and energy
conversion efficiencies. Biological conversion rates
are currently lower than chemical conversion
rates, and when considered alongside the chal-
lenges of potential product toxicity and a dilute
product output due to high water content, ad-
ditional robust bioprocess development is required
to reach metrics amenable for commercial scale
operation. This will require improvement in both
biocatalyst design and process development and
will rely on continued advancement in a number
of fields, including metabolic engineering and
synthetic and systems biology, among others.
These advancements are critical for the devel-
opment of bioconversion processes that can ex-
ploit the opportunity presented by currently
wasted, distributed methane feedstocks. Although
more mature bioconversion technologies target-
ing other feedstocks are currently available, no
other feedstock is as widespread and currently
available as methane. This further underscores
the ability to benefit from the learning-by-doing
approach to drive down CapEx costs and ad-
vance industrial biomanufacturing as a whole.
Given this potential, reducing time required for
biocatalyst and process development is essential
for industrial manufacturing to capitalize on this
current opportunity and advance the future of
the industry.
Strategies and tools within the iterative de-
sign, build, test, and learn cycle of biocatalyst
development remain a critical factor for unlock-
ing the potential of new approaches to biocon-
version. Expanding available metabolic pathways
through the development of novel synthetic en-
zymes and pathways to improve the carbon and
energy efficiency of conversion is critical to both
support and improve native pathways and orga-
nisms engineered for product synthesis. Aiding
these efforts are the use of in silico organism de-
sign strategies (71), genome mining techniques
(72), and computational enzyme design efforts
(73), in addition to exploiting emerging areas
such as substrate channeling (74). As opposed to
the traditional design of single microorganisms
for desired function, the use of microbial con-
sortia is an additional emerging approach that
Clomburg et al., Science 355, eaag0804 (2017) 6 January 2017
can potentially target applications challenging
for a single engineered organism (75). These
include engineering naturally occurring micro-
bial consortia while also developing synthetic
consortia to improve efficiency, stability, and
control. Beyond the traditional single-cell bio-
catalyst, another approach uses synthetic cell-
free systems, which convert a substrate to a
product using a mixture of enzymes and co-
factors for the cascade of reactions (76, 77). These
in vitro systems provide the potential for greater
control and flexibility by uncoupling product
synthesis from cellular viability, complexity, and
physiology and enabling high catalyst loading
to support high product titer and productivity.
Advancements in material design and three-
dimensional printing have the potential to im-
prove these efforts, with the use of printable
particulate methane monooxygenase-embedded
materials for conversion of methane to methanol
demonstrating the potential to improve not only
biocatalysts but also bioreactor and process
design (78).
Genome engineering technologies for making
mutations to microbial genes and genomes in a
targeted, multiplexed manner, such as CRISPR/
Cas9-based systems (7981) and multiplexed auto-
mated genome engineering (82), are becoming
standard tools for model industrial organisms.
The use of these tools allows in vivo construction
of the millions of potential biosynthetic pathway
variants generated by in silico metabolic meth-
ods and design in a fraction of the traditional
time. Furthermore, continued advances in syn-
thetic biology aimed at the development of de-
fined and standardized biological parts can be
exploited for the construction of minimal syn-
thetic genomes and organisms with desired
function (83). This type of modularization using
standard parts can improve the automation of
construction, serving to reduce both the time
and the costs associated with biocatalyst devel-
opment and improve upstream process design
efficiency.
The increasing throughput of design and con-
struction requires increasingly robust methods
for testing cellular functions that go beyond the
traditional analytical tools used today. Screening
and selection tools based on biosensors can help
increase throughput and decrease analysis time
(84). These techniques can be integrated with an
evolutionary approach to strain development,
with systems biology approaches enabling the
discovery of the genetic and physiological basis
of evolved phenotypes (85). Systems biology
tools, such as next-generation sequencing, high-
sensitivity proteomic and metabolomic methods,
and developments in fluxomic techniques, can
also be used to analyze and guide additional
rounds of rational design for further optimiza-
tion and biocatalyst performance refinement,
including improving the tolerance of biocata-
lysts to desired designer products (86).
Even with advancements in biocatalyst devel-
opment, a critical challenge is scaling production
from typical laboratory conditions used for strain
development to those required on a commercial
scale without the loss of performance. Typical
bioprocess development takes 5 to 10 years and
can be considerably more expensive than chem-
ical technologies owing to the relative infancy of
the process (87). The holistic consideration of
process parameters, including product purity
and waste-stream management, at an early stage
is desirable to develop small-scale testing pro-
tocols mimicking required larger-scale process
parameters. Modeling and designing novel bio-
reactors, especially those for gas-intensive fer-
mentations, will be critical to improving the
reproducibility and predictability of bioconver-
sion processes at the commercial scale. Process
intensification can build on initial success in
commercial-scale gas fermentation, with the
flexibility and modularity of biocatalyst devel-
opment enabling direct integration within the
Downloaded from http://science.sciencemag.org/ on February 16, 2017
process to shift product output. Early-stage as-
sessment of fermentation product separation
is also critical to success, because defining the
requirements for separation from a product, pro-
cess, and cost standpoint can help guide biocatalyst
development efforts and ensure full process in-
tegration. All of these process elements will ben-
efit from technological learning as bioconversion
processes are continually explored and devel-
oped, which will serve to reduce production and
energy costs and drive process optimization and
integration.
Targeting distributed methane feedstocks cur-
rently inaccessible through chemical processes
can help solidify industrial biomanufacturing
as an alternative to industrial chemical manu-
facturing and serve as an enabling factor for con-
tinued expansion and adoption for targeting a
range of complex feedstocks and products. Al-
though much work remains, the strengths and
diversity of biology can be leveraged to further
develop bioconversion processes for rapid, mo-
bile, and widespread deployment outside of those
previously demonstrated. This continued expan-
sion can further facilitate the ability for bio-
conversion to quickly adapt to market changes
or the arrival of new markets. Together, this
provides clear advantages for addressing the
emerging environmental, geographical, politi-
cal, and economic factors for industrial chem-
ical manufacturing. In addition to contributing
to solving complex challenges facing our planet,
the smaller scale and lower process complexity of
bioconversion processes also represent a promis-
ing and potentially transformative approach to
meet the demands of deep space exploration.
Extraterrestrial in situ resource use, product
manufacturing, and human health demands,
among others, will require innovative produc-
tion of manufacturing necessities using available
natural resources on location, such as atmo-
spheric gases (88, 89). Addressing the issue of
in situ resource use through biomanufacturing
solutions, given the potential for small-scale,
CapEx-efficient design, relative product flexi-
bility, and mild operating conditions will use
an innovative commercial technology in a novel
setting for potentially transformative solutions.
All told, the future of industrial biomanufacturing
8 of 10
R ES E A RC H | R E V IE W
holds great promise for the democratization of
chemical production and meeting the evolving
demands of chemical production in the 21st
century and beyond.
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AC KNOWLED GME NTS
This work was supported by grants from the U.S. National
Science Foundation (CBET-1605999) and the Advanced
Research Projects Agency-Energy of the U.S. Department of
Energy (ARPA-E DEAR0000762).
10.1126/science.aag0804
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 Industrial biomanufacturing: The future of chemical
production
James M. Clomburg, Anna M. Crumbley and Ramon Gonzalez
(January 5, 2017)
Science 355 (6320), . [doi: 10.1126/science.aag0804]

Editor's Summary

The next era of chemical manufacturing

Producing mass quantities of chemicals has its roots in the industrial revolution. But industrial
synthesis leads to sizeable sustainability and socioeconomic challenges. The rapid advances in
biotechnology suggest that biological manufacturing may soon be a feasible alternative, but can it
produce chemicals at scale? Clomburg et al. review the progress made in industrial biomanufacturing,
including the tradeoffs between highly tunable biocatalysts and units of scale. The biological

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conversion of single-carbon compounds such as methane, for example, has served as a testbed for more
sustainable, decentralized production of desirable compounds.
Science, this issue p. 10.1126/science.aag0804

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