J. Fish. Sci. Taiwan.

28(3): 151-162

Transglutaminase in Seafood and Meat Processings

Application of Transglutaminase in Seafood and Meat

Processings

Shann-Tzong Jiang and Li-Jung Yin

Department of Food Science, National Taiwan Ocean University, Keelung, 202, Taiwan.

Abstract
To search world-widely for protein sources and to broaden the application
potentials of existing proteins for human consumption have long been hot
topics for both food industry and scientists. Protein modification by enzymes,
especially by microbial transglutaminase (MTGase), the mass production of
which can be achieved by fermentation from cheap substrates, is one of the
most promising alternatives in developing novel proteins foods. MTGase, as
well as other TGases, can catalyze the formation of -(-Glutamyl)-Lysine
bonds in many food proteins. The resulting cross-link drastically alters protein
functionalities. TGase-catalyzed reactions can be used to modify the functional
properties of food proteins such as cross-linking of whey proteins, soya
proteins, gluten, myosin and actomyosin. The modification of food proteins by
TGase may lead to textured products, help to protect lysine in food proteins
from various chemical reactions, encapsulate the lipids and/or lipid-soluble
materials, form heat- and water-resistant films, avoid heat treatment for
gelation, improve elasticity and water holding capacity, modify solubility and
other functional properties, and produce food proteins with higher nutritive
value through cross-linking of different proteins containing complementary
limited essential amino acids. Accordingly, this review will focus on the
properties of MTGase, recombinant MTGase and other TGases. The
application of this enzyme in food processing will also be discussed.
Keywords: transglutaminase, recombinant transglutaminase, cross-linking of
proteins, seafood processing.

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Introduction
Transglutaminase (EC 2.3.2.13, TGase) is an enzyme which can catalyze acyltransfer reactions introducing covalent cross-link between proteins, peptides and
various primary amines by forming -(-Glutamyl)-Lysine bonds (G-L bonds) both
intra- and inter-molecularly (Ando et al., 1989; Folk, 1980; Ikura, 1988; Motoki and
Seguro, 1994; Nonaka et al., 1989). These TGase-catalyzed reactions can be used to
modify the functional properties of food proteins such as cross-linking of whey
proteins, soya proteins, gluten, myosin and actomyosin. The modification of food
proteins by TGase may lead to textured products, help to protect lysine in food
proteins from various chemical reactions, encapsulate the lipids and/or lipid-soluble
materials, form heat- and water-resistant films, avoid heat treatment for gelation,
improve elasticity and water holding capacity, modify solubility and other functional
properties, and produce food proteins of higher nutritive value through cross-linking of
different proteins containing complementary limiting essential amino acids (Motoki
and Seguro, 1994). To further understand the MTGase and TGase, this review will
focus on their properties and applications in seafood and meat processings.
Distribution of Transglutaminase
TGase is widely distributed in various tissues and body fluids (Folk, 1970; Hanigan
and Goldsmith, 1978) and also in plants (Folk 1980; Icekson and Apelbaum, 1987;
Yasueda et al. 1995), fish (Araki and Seki, 1993), pig blood (Jiang and Lee, 1992) and
microorganisms (Ando et al., 1989; Klein et al., 1992; Tsai et al., 1996a, b). It has been
purified from liver (Connellan et al., 1971) and hair follicle (Chung and Folk, 1972) of
guinea pig, rabbit liver (Abe et al., 1977), epidermal and erythrocyte (Brenner and Wold,
1978) of human, walleye pollack liver (Kumazawa et al., 1996), Japanese oyster
(Kumazawa et al., 1997) and also isolated from muscle and surimi of Alaska pollack
(Seki et al., 1990). TGases are also involved in several biological phenomena, such as
blood clotting, wound healing, epidermal keratinization, and stiffening of the
erythrocyte membrane (Aeschlimann and Paulsson, 1994). They are found to be
responsible for the regulation of cellular growth, differentiation, and proliferation.
Since the 1960s, the purification, characterization and application of Ca2+-dependent
TGase of animal origin, mainly guinea-pig liver, have been intensively studied
(Connellan et al., 1971; Folk, 1980; Ikura, 1988). It is noted that the setting
phenomenon of salted and ground fish flesh, so-called suwari, in the Japanese
kamaboko manufacturing may be closely associated with TGase-catalysed protein
crosslinking. The contribution of intrinsic TGase in fish to the suwari phenomenon has
been well demonstrated by Seki et al. (1990). TGase is believed to modify functional
properties of protein substrates.
In an attempt to develop new foods with unique proteins and processing
methodologies, modification of the functional properties through the formation of G-L
bonds by the catalytic action of TGase has been extensively studied.
Feasibility studies on protein modification by TGase
In the early 1980s, the possibility of modification of functional properties in milk
caseins and soybean globulins was demonstrated using TGase derived from guinea
pig liver (Ikura et al., 1989) or bovine plasma (Kurth and Rogers, 1984). In both
studies, cross-linking of food proteins of different origins and incorporation of amino
acids or peptides into food proteins to correct nutritive deficiency were shown. The
feasibility of food protein modification for industrial utilization using the guinea pig
enzyme was investigated by Motoki et al. (1989) and Motoki and Seguro (1994). In
these studies, they found that highly concentrated solutions of whey proteins, or
actomyosin from beef, pork, chicken or fish could be used as substrates and gelled by
the enzyme. Subsequently, improvement in solubility, water-holding capacity and
thermal stability of food proteins was demonstrated. Among these experiments, it also
found that protein solution mixed with the TGase could form a transparent and
water-resistant protein film. All these results indicated that TGase was potentially
useful in creating proteins with new unique functional properties. However, limited
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supply and unacceptability of guinea pig liver for food use greatly discouraged
commercialization. Mass production of this enzyme was probably the most essential
issue.
Production of microbial transglutaminase
Guinea-pig liver has been the sole source of commercial TGase for decades. The
limited source and complicated separation procedures resulted in increase in the cost
for obtaining this enzyme, which consequently makes it not possible to apply in food
processing. Recently, efforts have been made to search for TGase from
microorganisms (MTGase). MTGase was found in cultures of Streptoverticillium sp.,
Streptomyces sp. and Streptoverticillium ladakanum (Motoki et al., 1989; Ando et al.,
1989; Tsai et al., 1996a, b; Zeng et al., 2001). Microbial fermentation makes it
possible to achieve mass production of MTGase using cheap substrates. A number of
examples of the application of MTGase in food processing have been announced.
However, the potential for using MTGase in cosmetics, pharmaceutical products and
medical treatment, remains uncertain for commercial reasons and communication
difficulties.
Some MTGase producing microorganisms were screened using the hydroxamate
assay and taxonomically classified as a variant of Streptoverticillium mobaraense
(Ando et al., 1989; Washizu et al., 1994). These microorganisms excreted MTGase
into the culture broth. The critical property of MTGase, ability of the formation of G-L
bonds among proteins demonstrated that the produced enzyme was MTGase
(Nonaka et al., 1989). Motoki et al. (1989) reported that other Streptoverticillium
strains, such as St. griseocarneum, and St. cinnamoneum subsp. cinnanoneum also
have the ability to produce MTGase. It was also found in Streptomyces sp. (Ando et
al., 1989) and St. ladakanum (Tsai et al., 1996a, b).
The fermentation procedure for the production of MTGase is generally the same as
those mentioned microorganisms (Ando et al., 1989; Motoki et al., 1989). Glucose,
sucrose, starch, glycerine and dextrin can be used as carbon source. Inorganic as
well as organic nitrogen sources can be used, such as NH4NO3, (NH4)2SO4, urea,
NaNO3, NH4Cl, soya, rice, maize, wheat or wheat flour, bran, defatted soya bean,
maize-steep liquid, peptone, meat extract, casein, amino acids and yeast extract.
Necessary minerals and trace elements are phosphate, magnesium, potassium, iron,
copper, zinc and vitamins, etc. Non-ion surfactant and antifoam can also be added if
necessary. The culture is an aerobic fermentation so that aeration and agitation are
necessary. The temperature for growth and product formation is between 25C and
35C, and the fermentation time is normally 2-4 days (Sakamoto et al., 1992). Since
MTGase is excreted into the culture medium, cell disruption is unnecessary. Its
purification thus proves to be rather easy. Consequently, its commercialization has
been accelerated.
Examples of fermentation and purification of MTGase have been described by
Ando et al. (1989) and Motoki et al. (1989). The microorganism was activated at 30C
for 2 days and then cultured at the same temperature for 3 days under aeration (10
l/min) and agitation (250 rpm). The broth had an enzyme activity ranging from 0.28
U/ml to 2.5 U/ml, dependent upon the strain used. After being separated by
centrifugation at 3000 rpm, the supernatant was concentrated with an ultra-filtration
and then chromatographed on Amberlite CG-50 and Blue Sepharose. The total
recovery of MTGase activity was about 42%. However, recently a modified
downstream process for purifying MTGase was described by Ho et al. (2000). After
the fermentation of Streptoverticillium ladakanum, the broth was centrifuged and
filtered. The enzyme was purified directly with a rapid and simple stepwise
chromatography method with CM Sepharose CL-6B and Blue Sepharose Fast Flow.
According to the authors, this method is simple, rapid and has a MTGase recovery of
81%.

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Characteristics of MTGase
The properties of the enzyme have extensively examined (Connellan et al., 1971;
Abe et al., 1977; Folk, 1980; Lorand and Conrad, 1984; Kumazawa et al., 1996, 1997).
The endogenous TGase was found to be calcium dependent and with MW ranged from
77 to 90 kDa. The optimal temperature and pH were 35-50C and 7.5-9.0, respectively.
However, the MW of MTGase ranges from 30 to 45 kDa and it is calcium independent.
In general, the MW of TGase from animals is much higher than those from microbial
sources, such as: guinea pig liver TGase: 76.6 kDa (Folk et al., 1967; Ikura et al.,
1990); human erythrocyte TGase: 82 kDa (Brenner and Wold, 1978); human placenta
TGase: 76 kDa (De Backer-Royer and Meunier, 1992); rabbit liver TGase: 80 kDa
(Abe et al., 1977). The pI of animal-origin TGase has been found to be lower than 7.0,
such as: guinea pig liver TGase: 4.5 (Folk and Cole, 1966a, b; Folk et al., 1967);
rabbit liver TGase: 5.4 (Abe et al., 1977); human placenta TGase: 5.1(De
Backer-Royer and Meunier, 1992). It's optimal temperature and pH were 35-50C and
5.5-7.0, respectively. The MW of TGase from S. ladakanum was similar to that from
Streptoverticillium mobarense (MW: 40 kDa), while the pI (pI: 7.9)(Tsai et al., 1996a)
was lower than that from S. mobarense (pI 8.9) (Ando et al., 1989). The MTGase was
stable at pH 5.0~7.0 after 30 min incubation at 25C, in which more than 90% activity
was retained (Tsai et al., 1996a, b). This suggested that the MTGase was quite
resistant to variations of pH, compared with the narrow working pH of TGase from
animal sources (De Backer-Royer and Meunier, 1992). According to dynamic study on
MTGase from S. ladakanum, the rate constants for thermal inactivation at 50 and
55C were 6.2 x 10-4, and 2.1 x 10-3 sec-1, respectively (Tsai et al., 1996a), which were
higher than those from pig plasma (Jiang and Lee, 1992). The H* and Ea at 55C of
the MTGase were 33.6 and 34.3 kcal/mol, respectively (Tsai et al., 1996a), which
were lower than those from pig plasma (60.3 and 47.2 kcal/mol) (Jiang and Lee,
1992). These results suggested that the MTGase was less thermally stable and less
dependent on change in temperature than Factor XIIIa from pig plasma. Factor XIII
(pro-transglutaminase) was also purified from human plasma and platelet (Schwartz et
al., 1973). The catalytic properties of factor XIII were found to be different from those of
transglutaminase obtained from tissues (Chung and Folk, 1972) and microorganisms
(Ando et al., 1989; Tsai et al., 1996a, b; Jiang et al., 1998). However, because of the
limited amounts and difficulties of isolation and purification from animals, more
economical sources need to be found for the commercial utilization of TGase.
Physicochemical properties, such as molecular weight and secondary structures, and
enzymatic properties have already been reported (Ando et al., 1989; Nonaka et al.,
1989; Kanaji et al., 1993). Most of these data were obtained in the presence of
reducing agents for the purpose of comparison with the guinea pig liver enzyme.
However, reducing agents, dithiothreitol, -mercaptoethanol, and glutathione, would
noticeably change such properties as thermal stability and sensitivity to heavy metals.
Therefore, some of the properties were also determined in the absence of reducing
agents (Seguro et al., 1996).
The isoelectric point of MTGase was approximately 8.9. The molecular weight was
about 38,000. Protein sequencing by the automated Edman method and mass
spectrometry also revealed the primary structure of MTGase, comprising of 331
amino acid residues (Kanaji et al., 1993). The results from cDNA sequencing of the
MTGase gene coincided well with the result of protein sequencing. It further
suggested that MTGase possibly has a signal peptide of 18 amino acid residues at its
amino terminal (Washizu et al., 1994). The overall sequence data indicate that
MTGase has a single cysteine residue. The molecular weight calculated from the
amino acid composition (331 residues) is 37,842, which is similar to the
experimentally obtained value of 38,000. MTGase is, therefore, considered to be a
monomeric and simple protein (not a glycoprotein or lipoprotein), although there are
two potential glycosylation sites (-Thr-Xcc-Asn-) in the primary structure.
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Enzymatic properties of MTGase

The pH optimum of MTGase was around 5 to 8. However, even at pH 4 or 9,
MTGase still expresses some enzymatic activity. MTGase is thus considered to be
stable over a wide pH range. The MTGase has an optimum temperature of 50C and
fully sustained its activity even at 50C for 10 min. However, it lost activity within a few
min heating at 70C. MTGase still expressed activity at 10C, and retained some
activity at temperatures just above the freezing-point. Concerning substrate specificity,
most food proteins such as legume globulins, wheat gluten, egg yolk and egg white
proteins, myosins, fibrins, milk caseins, -lactalbumin and -lactoglobulin, as well as
many other albumins, could be cross-linked by MTGase (Nonaka et al., 1989, 1990;
Seguro et al., 1996).
TGase including the well-characterized guinea pig liver enzyme require Ca2+ for
enzyme activity. However, MTGase from Streptoverticillium sp. are totally
independent of Ca2+ (Ando et al., 1989; Tsai et al., 1996a, b; Jiang et al., 1998). In this
aspect, MTGase is quite unique from other mammalian enzymes. Such a property is
very useful in the modification of functional properties of food proteins, because many
food proteins, such as milk caseins, soybean globulins and myosins, are susceptible
to Ca2+. They are easily precipitated in the presence of Ca2+ and become less
sensitive to MTGase. The sensitivity of MTGase toward other cations in the absence
of reducing agents has also been investigated (Seguro et al., 1996) and found that
Cu2+, Zn2+, Pb2+and Li+ significantly inhibited MTGase. Because metals such as Ca2+,
Zn2+ and Pb2+ bind the thiol group of the single cysteine residue, this strongly supports
the idea that the cysteine residue could be part of the active site of MTGase.
MTGase is capable of gelling concentrated solutions of proteins such as soybean
proteins, milk proteins, beef pork, chicken and fish gelatin and myosin (Nielsen, 1995;
Nonaka et al., 1990; Seguro et al., 1996; Zhu et al., 1995), in a similar way to the
guinea pig liver enzyme. The gelled soya globulins further harden after 15 min heating
at 100C, resulting in novel properties. Milk caseins, non-heat setting proteins, were
also gelled by MTGase without heating. Gelatin, a cold-setting protein, was also
gelled. However, in this case, the gelled gelatin no longer melted on heating at 100C.
One of the features of MTGase is that two or more different proteins can be covalently
conjugated to produce new proteins with novel functionalities. For instance,
conjugation of milk casein or soya globulins to egg ovomucin, one of the glycoproteins,
improved the emulsifying activity relative to the parent proteins. Casein-gelatin
conjugation by MTGase also yielded novel proteins with a high solubility at acidic pH
(Kato et al., 1991).
MTGase is capable of incorporating amino acids or peptides covalently into
proteins (Nonaka et al., 1994). This reaction can improve nutritive values of food or
feed proteins, because covalently incorporated amino acids or peptides behave like
amino acid residues in a protein. For instance, caseins and soya proteins, in which
methionine and lysine, respectively, are limiting factors, could be improved by such as
MTGase reaction. In practical applications, all common amino acids, except lysine,
should have their -carboxyl group either amidated, esterified or decarboxylated to
eliminate the negative charge on the -carboxyl group. On the other hand, lysine itself
is a good substrate of MTGase because its -amino group is a primary amine.
Cloning and expression of TGase gene
TGase genes from guinea pig liver, chicken erythrocyte, microorganisms,
humans prostate, croaker, salmon, etc. have been successfully expressed in E. coli,
yeast or Streptomyces lividan. However, none of these TGases has been
over-expressed and commercialized.
Streptomyces lividan expression system
Washizu et al. (1994) used the Streptoverticillium S-8112 genomic DNA as
template and FDEEKGF and KVKQGWP taken from the amino acid sequence of the
TGase from Streptoverticillium sp. to design degenerate primers. They successfully
155

cloned a TGase cDNA with 1221 bp. This TGase cDNA was constructed on pAKW1
plasmid and transformed to St. lividan. After 5 days cultivation at 30C, MTGase with
MW of 37,000 was detected in the broth. This phenomenon suggested that the TGase
could be expressed in this system and excreted into broth, however, the expression
level was not high enough to commercialized.
E. coli expression system
E. coli transformed with pTTG2-22, which was constructed with rsbTGase from
croaker, was activated at 32oC for 14 h in YT broth with ampicillin, and cultivated for
another 20 h. The intracellular TGase was isolated (Yokoyama et al., 1998). From the
mentioned expression systems, the TGase can be expressed intra- or extra-cellular
depending upon the choice of cloning and hosting cells. Comparison of the amino
acids sequences of recombinant TGase from chicken erythrocyte, mouse phagocyte,
guinea pig liver and human endodermis cells, high homology on the N-terminus
MAEELVLETCDL was observed. This amino acid sequence was found to be highly
related to the formation of cross-linking among proteins (Boonmark et al., 1992;
Nakanishi et al., 1991; Yasueda, et al., 1995).
Application of transglutaminase
The production of transglutaminase by microorganisms makes it possible to
apply this enzyme in a variety of food processes. In meat processing it is of great
interest to maximize the yield of marketable products. This includes development of
methods for re-structuring low-value cuts and trimmings to improve their appearance,
flavour and texture and to enhance market value. Re-structuring treatment usually
involves size reduction, reforming and binding. In such a treatment, transglutaminase
can have a very important function. Sakamoto and Soeda (1991) developed a method
for producing minced-meat producing minced-meat products containing
transglutaminase. Minced meat and other food ingredients are mixed with
transglutaminase, shaped, packed in pressure-resistant containers and retorted to
manufacture meat products such as hamburgers, meatballs, stuffed dumplings and
shao-mai (a typical Chinese food). The foods show improved elasticity, texture, taste
and flavour. Minced beef and pork, flour, onion, skim-milk powder and condiments
were mixed with water and microbial transglutaminase, packed with sauce in bags
and retorted to make raw hamburger. Similar methods for meat and meat products
treated with transglutaminase can be found in the literature (Seguro et al., 1996).
Applications of MTGase in food processing
As already mentioned, many food protein substrates of MTGase, were gelled upon
incubation with it. The characteristics of such a gelation procedure and the gels
formed are as follows;
proteins that are not gelled by heating can be gelled;
gels that normally melts at elevated temperature no longer melt after the MTGase
gelation;
protein in oil-in-water emulsions, even in the presence of sugars and/or sodium
chloride, can be gelled;
gel firmness increases after heating;
the gels can no longer be solubilized by detergents or denaturants.
These characteristics are easily applicable for film, thread, etc, indicating that the
enzyme is very useful in protein texturization (Motoki and Seguro, 1994). The
following examples illustrate some of the practical industrial uses of MTGase in food
processing.
Meat products
MTGase can produce restructured meat by binding meat pieces at temperatures
below 10C, overnight. Kuraishi et al. (1996) have developed the new meat-binding
system using MTGase and caseinate simultaneously. Caseinate, when treated with
MTGase, becomes viscous, and the viscous caseinate acts as a glue to hold different
foodstuffs together. The effect of combining MTGase from Streptoverticillium
ladakanum with ultraviolet irradiation on the gelation of minced mackerel was
156

investigated by Jiang et al. (1998). It only needed 20 min to finish the gelation process
for mackerel mince at 30C, comparing with those without UV irradiation (needed
about 12 h). Using this system a larger piece of restructured meat like beef (or pork)
steaks or fish fillets from their smaller pieces can be prepared (Chung et al., 2000).
Meat pieces, including minced meat, can be also bound together by MTGase without
salt (sodium chloride) and phosphates, resulting in healthy meat products. The
MTGase treatment shows a synergistic effect, when combined with salt and
phosphates. In addition, MTGase enables low-fat meat production, and the use of
crosslinked caseinate as a fat substitute in Bologna-type sausages has been
suggested.
Fish products
Seki et al. (1990) and Tsukamasa et al. (1993) found that endogenous fish TGase
caused suwari setting, hardening fish protein paste at low temperature through
crosslinking. TGase from walleye Pollack, fish for surimi, has been purified and
characterized by Kumazawa et al. (1996). There seems to be no doubt that both
endogenous fish TGase and exogenous MTGase enable to improve the functionality
of fish raw materials by increase in cross-linking (Seki, 1992; Jiang et al., 2000a, b).
However, there are still some arguments as to whether the endogenous fish TGase is
the only factor in suwari setting (Nowsad et al., 1993).
A combination of TGase injection and tumbling results in reduced loss during
thawing and cooking in frozen fish products. Kumazawa et al. (1996) determined the
G-L bond in several fish eggs, and suggested that endogenous TGase may correlate
with the texture of raw and processed egg products. It seems that TGase treatment
improves and maintains the texture-quality of fish products, which strictly depends on
the freshness of raw materials.
Dairy products
Many researchers have shown that milk casein, which has no capability of gel
formation even by heating, was a very good substrate for various TGases. It was
found that a heat-resistant firm gel was formed from casein in the TGase reaction. An
example is the use of MTGase in yogurt production. Yogurt is a milk gel formed by
acidic fermentation with lactic acid bacterium starter, but it may suffer from the
problems of serum separation with a change in temperatures or physical impacts. This
phenomenon can be solved by adding MTGase, because MTGase improves the
water holding capacity of the gel. The MTGase reaction also makes it possible to
produce dairy products, such as ice cream and cheese with low-fat contents or a
reduced content of non-fat solids.
Soybean products
Soya proteins, such as 11s and 7s globulins, act as good substrates for the
MTGase reaction. Tofu, a typical soybean curd product, is prepared through
coagulation of soybean proteins with addition of Ca2+, Mg2+ and/or glucono--lactone.
It is very difficult to produce long-life tofu, since the soft, smooth texture of tofu is
easily destroyed by retort sterilization. However, the addition of MTGase enabled the
maintenance of the smooth texture of retorted tofu for a long time. MTGase was able
to improve the strength of yuba edible soybean film, a traditional delicatessen food
material in Japan.
Wheat products
Sakamoto (1996) found that MTGase treatment of noodles and pasta prevented
deterioration in textures after cooking, and improved the strength of the product, even
when low-grade flours were used. It was also suggested that the loaf volume of
several breads be increased or maintained by the addition of MTGase, when some
ingredients were substituted or reduced during mixing dough.
Other products (Motoki et al., 1989)
It is noteworthy that MTGase is able to introduce lots of new functions in gelatin and
collagen, in the way that their melting points and gel-strength are controlled freely with
the amount of MTGase. The MTGase treatment of gelatin resulted in heat-resistant
157

materials very similar to natural shark fins, a valuable food material in Chinese cuisine.
It is possible that sausage casings, presently prepared from collagen and chemical
reagents, could be replaced efficiently by the MTGase cross-linking of
collagen/gelatin or other proteins with no use of chemical reagents. In addition,
Watanabe et al. (1994) reported that the MTGase treatment reduced allergenicity of
wheat flours by modifying epitopic structures or opposing steric hindrance around the
epitopes. The MTGase treatment would be also one of solutions for patients suffering
from food allergy.
Perspectives
It is of paramount interest to search world-widely for protein sources and to
broaden the application potentials of existing proteins for human consumption. In
developing countries, many people are still suffering from starvation and efforts are
being focused on producing acceptable protein foods from non-animal proteins, to
solve the problem of protein deficiencies. On the other hand, in addition to their
awareness of health problems caused by obesity people in developed countries are
increasingly aware of the environmental burden caused by surplus livestock. Facing a
novel food product, consumers are very sensitive to properties such as flavour,
nutritional value, appearance, shelf life and palatability. In this respect, protein
modification by enzymes, especially by MTGase, the mass production of which can
be achieved by fermentation from cheap substrates, is one of the most promising
alternatives in developing novel proteins foods. MTGase, as well as other TGases,
can catalyze the formation of the G-L bond in many food proteins. The resulting
cross-link drastically alters protein functionalities. Applications are emerging in the
development of novel foods and non-food processing methods. There may be many
applications in the incorporation of various amines, amino acids, lysine-containing
peptides, glutamine-containing peptides and heterologous polypeptides. There is no
doubt that MTGase technology will be an essential tool for protein modification in both
food processing and non-food processing in the future. TGase-catalyzed reactions
can be used to modify the functional properties of food proteins such as cross-linking
of whey proteins, soya proteins, gluten, myosin and actomyosin. The modification of
food proteins by TGase may lead to textured products, help to protect lysine in food
proteins from various chemical reactions, encapsulate the lipids and/or lipid-soluble
materials, form heat- and water-resistant films, avoid heat treatment for gelation,
improve elasticity and water holding capacity, modify solubility and other functional
properties, and produce food proteins with higher nutritive value through cross-linking
of different proteins containing complementary limited essential amino acids.
Although guinea pig liver has been the sole source of commercial TGase for decades,
the limited source and complicated separation procedures resulted in increase in the
cost for obtaining this enzyme and consequently made it not possible to apply in food
processing. Microbial fermentation can achieve mass production of MTGase using
cheap substrates. With respect to the production of MTGase, the microbial process
has no doubt an advantage in its independence from regional and climatic conditions,
in addition to its reasonable cost. But it is still of great interest to improve fermentation
and down-stream processing to reduce production cost and waste further. In addition,
the production of MTGase, St. mobarense and St. ladakanum need 2 days to activate
and 4 days to ferment. This long fermentation period makes the cost much
increased.
Modification of strains by genetic engineering would be one of the alternatives.
The study on the cloning of MTGase gene from Streptoverticillium sp., expressing the
reconstructed gene in Escherichia coli and/or Pichia pastoris expression systems,
using site-specific mutation, Error Prone PCR or DNA Shuffling techniques to improve
the properties and finally applying the recombinant MTGase in meat and vegetarian
food processing would be the main topic on this area.

158

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161

~ (Microbial
Transglutaminase, EC 2.3.2.13, MTGase)
MTGase (TGase)
(glutamine
residue)-carboxyamide group -amino group
- (-glutamyl)lysine TGase

TGase

MTGase MTGase TGase

162