Saliva: composition and function

Em-on Benjavongkulchai
Department of Biochemistry

Objectives
1.Explain technical terms in saliva field
2.List components in saliva
3.Explain functions and roles of each
component
4.List influential factors of salivary flow rate and
composition.

References
Edgar WM and O'Mullane DM. Saliva and
Oral Health, 1996, Thanet Press Ltd,
Margate, Great Britain.
Tenovuo JO. Human Saliva: Clinical
Chemistry and Microbiology. vol. I & II,
1989, CRC Press Inc., Boca Raton,
United States.

Keyes rings surrounded by a saliva ring

Saliva

Host

caries

substrate

microflora

Host (Tooth)
Age
Genetics
Morphology
Microporosity
Apatite carbonate
Fluoride
Trace elements
Nutrition

Substrate
Carbohydrate
Food detergency
Eating frequency
Food physical
characteristics
Oral clearance

Saliva
Host

Substrate

Flora

Saliva
Bicarbonate level
Buffer capacity
Composition
Flow rate
pH

Flora
Streptococci
Lactobacilli
Virulence factors
Transmissibility
Fluoride in plaque
Oral hygiene

Properties of human saliva

pH

6.5-7.5

Specific gravity

1.01-1.02

Viscosity (shear rate 450 sec)

0.9-4 Centipoise

Lubrication

637 %Glycerol

Depression of freezing point

0.34-0.70 C

Ash

0.2

Dry substance

0.1-1.0 g%

Flow rate: Unstimulated

0.1-0.8 ml/min

Stimulated
Total volume

g%

1.1-3.0 ml/min
1-1.5 l/d

Contributions of the different salivary

glands to total daily secretion of saliva
Salivary gland

% of total saliva

parotid

25

submandibular

71

sublingual

3-4

minor (labial, palate, buccal) traces

Definition / Nomenclature
Saliva : mixture of secretions in the oral cavity
whole saliva (mixed saliva), oral fluid
parotid saliva :- parotid cup
submandibular saliva

submandibular / sublingual saliva

minor salivary gland secretions :microcapillary tube, filter paper
gingival crevicular fluid :microcapillary tube, filter paper

unstimulated saliva: no stimulants

resting saliva: ideal (not appropriate)

stimulated saliva :1. gustatory stimuli : 2% citric acid

2. masticatory stimuli : paraffin wax,
parafilm, neutral gum base,
rubber band

Source of whole saliva

parotid secretion
submandibular secretion
sublingual secretion
minor salivary gland secretion
lingual (von Ebner) secretion
gingival crevicular fluid (GCF)
microbiota
microbial enzymes
sloughed degeneration oral epithelial cells
food debris

Saliva secretion: Two stage hypothesis

JH Thaysen
First stage: production of isotonic primary saliva
Second stage: reabsorption of Na+ and Cl- results
in hypotonic secondary saliva
Myoepithelial
cell

Venous
blood
Striated Na+ Cl- Excretory
duct
duct

Saliva

1
Na+ Cl-

Acinar
cell

Intercalated
duct

Arterial
blood

Functions and Roles of Saliva

protective function
- lubrication
- physical cleansing
- chemical buffering
- antimicrobial activity
- remineralization
alimentary function
- tasting
- chewing
- digesting
- swallowing

Composition
water
Inorganic compounds

Organic compounds
Proteins
Lipids
Sugars

Organic compounds

Amino acids
Metabolites
Hormones
Vitamins
Drugs, toxic substances

Sodium Na
+
Potassium K
2+
Calcium Ca
2+
Magnesium Mg
Chloride Cl
3Phosphate PO4
Bicarbonate HCO3
Fluoride F
Thiocyanate SCN
Hypothiocyanite OSCN

Copper
Cobalt
Lead
Strontium
Iodide
Bromide
Nitrate
Nitrite
Sulfate

Na , Cl : osmolarity of saliva
hypo-, iso-tonic
2+

3PO4 :

Ca ,
remineralization
2+
3supersaturation of Ca , PO4
HCO3 :

pH/buffer control

F : remineralization
Fluorapatite, CaF2
-

SCN /OSCN :
antimicrobial/detoxification
function

HCO3 :
pH

pH/buffer

7
Saliva

6
5

Saliva
-HCO3

4
H2O

1.0

2.0
0.02 M HCl

ml

H2CO3

HCO3

HCO3

2CO3

H2PO4

+H

pKa = 6.38

pKa = 10.32

+H

2HPO4

+H

pKa = 7.12

Phosphate
COO
Protein

+
NH4

O
Urea

H2N

NH2

Electrolyte concentration (mM) of human

submandibular saliva compared with plasma

Na
+
K
Cl
HCO3
2+
Mg
2+
Ca
Pi
pH

Saliva
Unstimulated*
2.6
14.4
11.9
2.2
0.07
1.56
3.6
6.47

* 0.26 ml/min, ** 3.0 ml/min

Plasma
Stimulated**
54.8
13.7
32.3
35.3
0.04
2.13
1.57
7.62

143.3
4.1
100.9
27.5
0.92
1.24
0.37
7.4

Electrolyte concentration (mM) of human parotid

saliva compared with plasma

Na
+
K
Cl
HCO3
2+
Mg
2+
Ca
Pi
pH
Osmolality
(mOsm/kg)

Saliva
Unstimulated*
2.7
46.3
31.5
0.6
0.18
1.04
3.35
5.82
85.7

*<0.011 ml/min, ** >1.0 ml/min

Plasma

Stimulated**
63.3
18.7
35.9
29.7
0.02
0.94
1.02
7.67
132.0

143.3
4.1
100.9
27.5
0.92
1.24
0.37
7.4
296.0

Factors influencing salivary composition

Species, Gender, Age

Hormones

Glandular source

Pregnancy

Flow rate

Genetic polymorphisms

Duration of stimulation

Stop-flow conditions

Previous stimulation

Antigenic stimulation

Circadian rhythms

Exercise

Nature of stimulus

Drugs

Plasma Composition - Diet

Various Diseases

Parotid
saliva

concentration
mM 60

Na

Cl

40

HCO3

20

P 2+
Ca 2+
2
Mg x10
0.5

1.0

1.5

Flow rate ml/min

320

200

0.12
80
0.06
sleep

sleep
awake

sleep

awake

sleep
awake

awake

0.00
12.00

12.00

12.00

12.00

12.00

Effect of circadian rhythm and lack of sleep on the

protein concentration of unstimulated parotid saliva

Unstimulated flow rate ml/min

Protein concentration mg/100 ml

Circadian rhythms

Nature of stimulus
Ascorbic acid and NaCl concentration (g%)
Salivary flow rate (ml/min)

10

0.50

15

20

Ascorbic acid
NaCl

0.25

Sucrose

40

80

Sucrose concentration (g%)

Protein synthesis
and secretion

ribosome
mRNA
growing polypeptide chain

Protein inside endoplasmic

reticulum
rough
endoplasmic
reticulum

glycosylation

Golgi apparatus

secretion vesicle

exocytosis

Plasma membrane

-Amylase

AD 1814: -amylase in malt

AD 1826: -amylase in human saliva
ptyalin
serous cells is a major
source of -amylase

-Amylase

CH2OH

CH2OH

O
O

-Amylase

m
CH2

CH2OH

O
O

Starch

Maltose,
maltotriose,
dextrins

-1,4-glycosidic bond

endoglycosidase

Characteristics:
2+

metalloenzyme :- Ca
pI 5.7-6.8
pH optimum 7.0

Roles:
Starch/glycogen digestion
Tissue coating: binding to hydroxyapatite
Binding to some oral bacteria eg. Streptococcus
gordonii, S mitis, not S sanguinis, S oralis
Antimicrobial agent: inhibit growth of Neisseria
gonorrhoeae, Legionella pneumophila

Salivary peroxidase
serous cells
submandibular gland
parotid gland
sublingual gland

Salivary
peroxidase

Peroxidase / thiocyanate (SCN ) /

hypothiocyanite (OSCN )
salivary peroxidase

SCN + H2O2
thiocyanate

OSCN + H2O2

OSCN + H2O
hypothiocyanite

salivary peroxidase

SCN + O2 + H2O

Salivary
peroxidase

Toxicity of H2O2
oxidising agent

.OH

hydroxyl
radical

-2

.O

superoxide
radical

H2O2
hydrogen
peroxide

High

O2

dioxygen

Low
Toxicity

Salivary
peroxidase

H2O2
oxidise lipid, protein, DNA
toxic to bacteria & mammalian cell
2+

H2O2 + Fe

3+

Fe

+ .OH + OH

Salivary
peroxidase

Production of H2O2
Streptococcus mitis
Streptococcus sanguinis
pyruvate oxidase

pyruvate + O2 + Pi
+

H2O2 + CO2 + acetyl

phosphate
NADH oxidase

NADH + H + O2

H2O2 + NAD

Myeloperoxidase from leukocyte

-

Cl + H2O2

myeloperoxidase

OCl + H2O
hypochlorite

Characteristics:
pI 6-9
pH optimum 5-7
Glycoprotein
Metalloenzyme Fe

Roles:

3+

Antimicrobial activity

OSCN inhibits bacterial glycolysis via

glyceraldehyde-3-phosphate dehydrogenase,
hexokinase

Protection of host cell from H2O2 toxicity

Lysozyme (Muramidase)
-1,4-Glycosidic linkage
CH2OH

CH2OH

O
O

R = lactyl side chain

OR

NH

NH
O

C
CH3

N-Acetylmuramate (NAM)

C
CH3

N-Acetylglucosamine (NAG)

Characteristics:
- pI = 11
- pH optimum = 5
- Glycoprotein

Role:
Lysis of peptidoglycan
Activation of bacterial autolysin
Aggregation of bacteria
Inhibition of bacterial adhesion to tooth surfaces
Inhibition of glucose uptake and acid production

Lactoferrin
Characteristics

- Glycoprotein
- bind 2 atoms of iron/molecule
- synthesized from glandular
epithelial cells & neutrophils

Roles
- Deprivation of iron from bacteria
(bacteriostatic action) Streptococcus mutans
Candida albicans
- Bactericidal action by direct contact
of iron-free form of lactoferrin (apo-LF)
- Inhibition of bacterial adhesion to
tooth surfaces

- Aggregation of bacteria

Statherin
Characteristics
- MW 5,380 dalton
- pI 4.2

- glycoprotein
- phosphoprotein 2 P/molecule

- synthesized from serous cell

- tear fluid, nasal & bronchial mucus

statherin

Roles
- inhibit calcium phosphate precipitation

- inhibit calcite precipitation

- acquired pellicle formation

- bacteriostatic/bacteriocidal activity of
anaerobic oral bacteria

Bacteroides forsythus
Bacteroides ureolyticus
Fusobacterium necrogenes
Fusobacterium necrophorum

Proline rich proteins

synthesized from serous cell
phosphoprotein 2 P/molecule
binding to tannin
3 groups
1. acidic PRP (aPRP)
31% total parotid salivary protein
2. basic PRP (bPRP)
23%
3. glycosylated PRP (gPRP)
17%

Acidic PRP
Saliva specific protein
Roles
- adsorption to hydroxyapatite
- formation of acquired pellicle
- binding of ionic calcium
- inhibition of calcium phosphate
precipitation
- inhibition of apatitic crystal growth

Salivary cystatins
cysteine protease inhibitors
-serous cells of submandibular/parotid
-subman > > parotid
- cystatin S (saliva)
cystatin SN (neutral)
cystatin SA (acidic)

Cystatins

Roles
1. regulate protease activity eg. inflammation
process in gingivitis, periodontitis
2. acquired pellicle formation

3. inhibit hydroxyapatite crystal growth

Mucins
Synthesized from mucous acini
(submandibular, sublingual
glands, minor salivary glands)

MUC1, MUC4
MUC5B (MG1 = high molecular weight mucin)
MUC7 (MG2 = low molecular weight mucin)

Mucins

MGI (MUC5B)

MG2 (MUC7)

subunit

yes

no

protein content (%)

14.9

30.4

CHO content (%)

78.1

68.0

1.6

sulfate (%)
MW (kD)

1000

CHO-linkage

O-linked

fatty acids

5-10

150 - 180
O-/N- linked
0

Mucins

Roles
- Tissue coating and pellicle formation
- Lubrication

- Antimicrobial activity
- Substrate for microbial metabolism

Histatins
(histidine rich proteins)
Saliva-specific proteins
parotid, submandibular/sublingual
MW ~ 1-5 KD (7-38 amino acids)
pI 7-9
non-glycoprotein
metalloprotein, copper & zinc
10-150 g/ml (3-50 M)

Histatins

Roles & functions

acquired pellicle formation

inhibit calcium phosphate

precipitation

antibacterial activity, inhibit growth &

viability

antifungal activity, inhibit

Candida albicans
activate intrinsic autolysis
membrane damage

Carbonic anhydrase
- 1933 red blood cells

- serous cells
- liver, lung, kidney, muscle etc.

CO2 + H2O

H2CO3
carbonic
anhydrase

pKa = 6.4

HCO3

+ H

Carbonic
anhydrase

Functions & Roles

buffer capacity & pH regulation
Characteristics

single polypeptide chain

glycoprotein
MW 42 kD (deglycosylated form = 36 kD)
salivary carbonic anhydrase = isozyme II,
isozyme VI
2+

metalloenzyme, Zn

pH
8.0

mmHg mEq/l
44

pH
HCO3

60

pCO2
7.2

36

40

6.4

28

20

5.6

20

1.0

2.0

3.0

Flow rate ml/min

bicarbonate concentration and pH flow rate

Antimicrobial Agents in
Human Whole Saliva
Agglutinins*, aggregating factors
Anionic antimicrobial proteins
Amylase*
Basic proline-rich protein*
Cationic antimicrobial proteins
Histatins*, Defensins*, Cathelicidin
Immunoglobulins
Lactoferrin*
Lysozyme*
Mucins*
Myeloperoxidase system*
Salivary peroxidase system*
Statherin*
Cystatins

statherin agglutinin -amylase

-defensin lysozyme cystatins
histatins
peroxidases mucins
carbonic anhydrases
lactoferrin
histatins
antiPRPs
mucins
bacteria buffering
cystatins
-amylase
peroxidases
lactoferrin
anti-viral
digestion mucins
lysozyme

histatins
cystatins
mineraliza
lactoferrin anti-fungal
histatins
tion
lysozyme
statherin
-defensin
tissue
lubrica
PRPs
coating
tion
-amylase
histatins
mucins
PRPs
mucins
statherin
statherin
cystatins

References
Duckworth RM. The teeth and their
environment: Physical, chemical and
biochemical influences. Monographs in Oral
Sciences. Vol 19, 2006.

Fejerskov O, Kidd E. Dental caries: The disease

and its clinical management. 2003, Blackwell
Munksgaard, Oxford.
Newbrun E. Cariology. 1989. 3rd ed.
Quintessence Publishing, Chicago.
Wefel JS, Donly KJ. Cariology. Dent Clin North
Am. 1999; 43:569-780.

Objectives
The students should be able to
1. explain definition, formation, composition and
functions of acquired enamel pellicle
2. discuss roles of acquired pellicle in dental
caries process.

Film on hard surface,

tooth and prosthesis
Film on tooth:

acquired enamel pellicle

Film on soft tissue:

Mucosa pellicle

Definition
acellular layer of adsorbed salivary
proteins and other macromolecules on
the dental enamel surface
< 1 m

A thin heterogeneous film of

biomaterial

TEM micrographs of pellicle layer formed in situ on enamel specimens

mounted at the buccal surface of the upper first molar
1 min

10 min

60 min

120 min

200 nm

Formation
Selective adsorption of proteins
Interaction between salivary

proteins and tooth

surface/hydroxyapatite

Formation
Interaction between

positively charge ion from

saliva and phosphate ion from
hydroxyapatite

Hydration layer (+ve)

Ca: Phosphate = 10:1
attract

Proteins

Adsorption of salivary proteins to enamel surface

via various electrostatic interaction

Interaction between
salivary proteins and tooth
surface/hydroxyapatite
Weak/reversible
Physicochemical interaction
Electrostatic interaction
Van der Waals
H-bonding

Binding of proteins to enamel

2 steps
rapid adsorption (direct binding)
slow adsorption (protein-protein

interactions)

Proteins with high affinity

binding to hydroxyapatite
Group 1 Fast PRP3, PRP4, statherin
Group 2 -amylase, glycosylated PRP,

cystatin
Group 3 PRP1, PRP2, histatins

H2PO3 H2PO3
5

10

NH 3-ASP-SER-SER-GLU-GLU-LYS-PHE-LEU-ARG-ARG-ILE-GLYARG-PHE- - - - - +
+ +
+
15

20

25

GLY-TYR-GLY-TYR-GLY-PRO-TYR-GLN-PRO-VAL-PRO-GLU-GLNPRO-LEU30

35

40

43

TYR-PRO-GLN-PRO-TYR-GLN-PRO-GLN-TYR-GLN-GLN-TYR-THRPHE-COO

Primary structure of human statherin

Primary Structure of Human

Acidic Proline-Rich Protein 1
P
pGLU-ASP-LEU-ASP-GLU-ASP-VAL-SER-GLN-GLU-ASP-VAL-PRO-LUE-VAL-ILE-SER-ASP-GLY-GLY

- -

P
ASP-SER-GLU-GLN-PHE-ILE-ASP-GLU-GLU-ARG-GLN-GLY-PRO-PRO-LEU-GLY-GLY-GLN-GLN-SER

- -

GLN-PRO-SER-ALA-GLY-ASP-GLY-ASN-GLN-ASN-ASP-GLY-PRO-GLN-GLN-GLY-PRO-PRO-GLN-GLN

GLY-GLY -GLN-GLN GLN-GLN-GLY-PRO-PRO-PRO-PRO-GLN- GLY-LYS-PRO-GLN-GLY-PRO-PRO-GLN

GLN- GLY- GLY-HIS-PRO-PRO-PRO-PRO-GLN-GLY-ARG-PRO-GLN-GLY-PRO-PRO-GLN-GLN-GLY-GLY

HIS-PRO-ARG-PRO-PRO-ARG-GLY-ARG-PRO-GLN-GLY-PRO-PRO-GLY-GLN-GLY-GLY-HIS-GLN-GLN
+
+
+
+
130
GLY-PRO-PRO-PRO-PRO-PRO-PRO-GLY-LYS-PRO-GLN-GLY-PRO-PRO-PRO-GLN-GLY-GLY-ARG-PRO
+
+
150
GLN-GLY-PRO-PRO-GLN-GLY-GLN-SER-PRO-GLN

Interaction between salivary proteins and

other solid surfaces
Titanium

Titanium oxide TiO2

2+

Ca

2+

Ca
2+
Ca

amalgam
hydroxyapatite
titanium
glass
plastic
bacterial cell
mucosa
stainless steel
monocrystalline sapphire
polycrystalline alumina
siliconized germanium

Source of pellicle
composition
Saliva

Gingival Crevicular Fluid (Plasma)

Bacteria

Composition
Proteins
Glycoproteins,
phosphoproteins, enzymes

Plasma proteins
Salivary proteins
Bacterial proteins

Proteins and other biopolymers in pellicle layer

formed in situ/in vivo on enamel or hydroxyapatite
Biopolymer

Physiological role

Molecular weight (kDa)

Proline-rich proteins

pellicle precursor

5-30

Statherin

inhibits precipitation of calcium

Histatin

anti- Candida species

3-5

Glucosyltransferase

bacterial enzyme

140

Albumin

carrier protein, regulation

of colloid-osmotic pressure

69

sIgA

immune response, inhibition

of bacterial attachment

60/90

Fibrinogen

factor I of blood coagulation

340

Cytokeratin 13, 15

cell protein

40-67

Salivary agglutinin

agglutination of oral bacteria

300-400

Lipids
22-23% of dry weight
Neutral lipids: Free fatty acid Triglycerides,
Cholesterol, Cholesteryl esters
Phospholipids: Phosphatidyl
ethanolamine, Sphingomyelin,
Phosphatidyl choline

Glycolipids: Neutral and sulphated

glyceroglucolipids

Carbohydrates
Glucose
Galactose
Mannose
Fucose
Glucosamine
Galactosamine
Glucan

Functions & Roles

Lubrication of the tooth surface
Protecting the enamel against
abrasion and attrition

Semi-permeable barrier
Permeable-selective nature
Control ions in and out

of the tooth
Control de-/re-mineralization

Mineral homeostasis
No pellicle
Calcium phosphate precipitation
on enamel surface

With pellicle
Inhibition of calcium phosphate
precipitation on enamel surface

Modulation of bacterial adherence

Bacterium
Streptococcus mutans

Binding sites in pellicle

Glucan, Acidic proline-rich proteins, High MW mucins

Streptococcus sanguinis Complex of secretory IgA light chain and -amylase,

-Amylase, Fibrinogen, Lysozyme, Salivary mucin (MG2)
Streptococcus gordonii

Acidic proline-rich proteins, -Amylase, Fibrinogen,

Salivary mucin (MG2), Glucan

Streptococcus pyogenes Glucan

Streptococcus mitis

Fibrinogen, -Amylase, Glucan

Streptococcus oralis

Fibrinogen, Salivary mucin (MG2), -Amylase

Streptococcus sobrinus

Glucan

a complex soft non-calcified

microbial deposit which
accumulates on the surfaces of
the teeth

References
Duckworth RM. The teeth and their
environment: Physical, chemical and
biochemical influences. Monographs in Oral
Sciences. Vol 19, 2006.

Fejerskov O, Kidd E. Dental caries: The disease

and its clinical management. 2003, Blackwell
Munksgaard, Oxford.
Newbrun E. Cariology. 1989. 3rd ed.
Quintessence Publishing, Chicago.
Wefel JS, Donly KJ. Cariology. Dent Clin North
Am. 1999; 43:569-780.

Objectives
The students should be able to
1. explain definition, formation,
composition and functions of plaque, and
calculus
2. discuss roles of plaque and calculus in
dental caries process.

Acquired pellicle

Biofilm formation

Formation

Interaction between oral bacteria

and salivary proteins
Cell surface, Fimbriae, Pili

Interaction between oral bacteria

and salivary proteins

Physicochemical interaction
Electrostatic
Weak
Hydrophobic
interaction
Van der Waals
H-bonding
Specific binding
Protein receptors

Strong
interaction

Salivary protein
Hydrophobic
interaction
Carbohydrate
antigen

Lipoteichoic
acid (LTA)

Fimbriae
Cell wall
Cell membrane
Bacterial cell

Strong interaction
Bacterial Antigen I/II
(spaP, sspA, sspB etc.)

Salivary
Agglutinin

H2PO3 H2PO3
5

10

NH 3-ASP-SER-SER-GLU-GLU-LYS-PHE-LEU-ARG-ARG-ILE-GLY-ARG-PHE- - - - - +
+ +
+
15

20

25

GLY-TYR-GLY-TYR-GLY-PRO-TYR-GLN-PRO-VAL-PRO-GLU-GLN-PRO-LEU30

35

40

43

TYR-PRO-GLN-PRO-TYR-GLN-PRO-GLN-TYR-GLN-GLN-TYR-THR-PHE-COO

Actinomyces naeslundii gsp2

Fusobacterium nucleatum

Amino acid sequence of human statherin

Binding (%)

(a) A viscosus R28 (type-I)

Statherin

80

S33-38
S39-43

60

S27-32
40

20

S21-26
S16-20
0

10

15

20

Concentration (M)

Type 1 Fimbriae A naeslundii Proline-rich proteins

Antigen I/II (P1) S mutans

Agglutinin

A naeslundii
Fusobacterium nucleatum

Statherin

S gordonii

-amylase

Primary colonizers
S. sanguinis, S. oralis,

S. mitis, Actinomyces spp

Late colonizers
S. mutans,
Porphyromonas gingivalis,
Fusobacterium nucleatum

Streptococcus sanguinis
Streptococcus oralis
Terminal sialic acid residues in salivary
glycoproteins

Streptococcus oralis
Galactose binding adhesin

Acquired pellicle
Biofilm formation

Coaggregation
Direct cell-cell binding
Bridging of cells by salivary proteins
Adhesins: cell wall, fimbria, pili
Multiple adhesins

Adhesins and binding proteins of human oral bacteria

Adhesin/binding Subunit size
Protein
Gram-positive
SspA

Fibrils (91 nm)

(kDa)

Bacterial
strain

Inhibitor

(molecule/cell)

210 [AgI/II] S gordonii DL1

A naeslundii

170 [AgI/II] S gordonii G9B

P gingivalis 33277

100
fimbriae

S gordonii DL1
S oralis 34
A naeslundii T14V GalNAc(13)

Gram-negative
PlaA

Receptor

40
42

GalNAc
Gal

Gal

Gal

Gal(13)

GalNAc

P gingivalis 33277 A naeslundii WVU627 F nucleatum T18

P gingivalis T22

Lactose

Adhesins and binding proteins of human oral bacteria

Adhesin/binding Subunit size Bacterial
Protein

(kDa)

strain

Receptor

Inhibitor

(molecule/cell)

Lipoproteins
Group1
SsaB
FimA

35
35

S sanguis 12
S parasanguis
FW213

Saliva
Saliva, fibrin

Group2
HppA
HppG

76
78

S gordonii DL1
S gordonii DL1

Hexa heptapeptide
Hexa heptapeptide

45

S mutans LT11

Mellibiose, raffinose

185
210

S mutans NG5
S sobrinus 6715

Salivary agglutinin
Salivary agglutinin

Group3
MsmE
Antigen I/II
SpaP
SpaA

Other mechanisms
in plaque formation

Glucosyltransferases
Fructosyltransferase
Glucan binding proteins

Dextransucrase
Glucansucrase

GTFs (Glucosyltransferases):
Streptococcus sp
Dextransucrases, alternansucrase:
Leuconostoc mesenteroides
GTFs: Lactococcus sp, Lactobacillus sp

Extracellular enzyme

Leuconostoc mesenteroides
Streptococcus bovis
Streptococcus mutans
S.sanguinis
S.salivarius
S. sobrinus etc.
Lactobacillus sp.
14 gtf genes

n glucose-fructose

(sucrose)

Glucosyltransferases
(GTFs)
glucosen + n fructose
(glucan, dextran,
mutan)

n glucose-fructose

(sucrose)

Fructosyltransferases
(FTFs)
fructosen + n glucose
(fructan, levan)

Dextran: Water soluble

-1,6- glucosidic linkage

Branch -1,2-; -1,3-; -1,4-

Mutan: Water insoluble

-1,6- glucosidic linkage
High degree of -1,3-

5
4

1
3

2
6

o
1

Glucan (dextran)

Portion showing principle 1:6

linkage and branching at C3 and C4

CH2
O

Fructan
(Levan)

Portion showing
principle 2:6
linkage &
branching at C1

Solubility: type and degree of branching

number of monomer
Levan -2,6-: Water soluble, Levan -2,1-: Water insoluble

GTFs and FTFs have very high

specificity to sucrose
Sucrose: A specific substrate for GTF
: Glucosyl Donor

UTP

glucose-1-P

UDP-glucose +

PPi

Glycogen synthesis

Free energy of hydrolysis of glucosyl donors

and some polysaccharides
Compound

Standard free energy of

hydrolysis GH(kcal/mole)

Glucose-1-phosphate

- 5.0

UDP-glucose *

- 7.6

Sucrose

- 6.6

Maltose

- 3.0

Lactose

- 3.0

Glycogen

- 4.3

Dextran

- 2.0

Levan

- 4.6

*UDP, uridine diphosphate

Km values (mM) for sucrose for GtfB, GtfC,

and GtfD in solution and when adsorbed
onto the surface of hydroxyapatite
Phase

GtfB

GtfC

Solution
Surface

30 + 4
12 + 4

20 + 5
10 + 1

GtfD

165 + 20
22 + 2

J Dent Res 1995, 74(10)

GBPs present among a variety of

streptococci
GbpA 1979

gbpA 1990

GbpB 1982

gbpB 2001

GbpC 1997

gbpC 1997

GbpD 2002

gbpD 2002

Glucan-binding Proteins among the Mutans Streptococci

Host
Protein Gene
S. mutans Ingbritt
GbpA gbpA
S. mutans SJ32
GbpB gbpB
S. mutans 109c
GbpC gbpC
S. mutans UA159
GbpD gbpD
S. sobrinus 6715-49 GBP-1 NC
S. sobrinus B13
GBP-2 NC
S. sobrinus 6715
GBP-3 NC
S. sobrinus 6715 GBP-4/GBL NC
S. sobrinus 6715
GBP-5 NC
S. sobrinus UAB108 Dei dei

Size(kDa)* Function
58
Biofilm morphology
41
Peptidoglycan hydrolase
58
Aggregation
75
Cohesion/enzyme
15-16 Unknown
71-87 Aggregation
52-68 Unknown
58-60 Aggregation
41-49 Unknown
31
Dextranase inhibitor

GbpA: biofilm architecture

GbpB: biofilm forming abilities
GbpC: biofilm thickness
GbpD: biofilm strength, lipase

Adhesion and
Colonization of Bacteria
Cell surface protein P1 (PAc, Ag I/II)

Fimbrillin
Insoluble/Soluble Glucan
Glucan Binding Protein

TEM of acquired pellicle and plaque formed

in situ within 24 h on enamel surface

Dental
plaque

Acquired
pellicle

Roles
Dental caries
Plaque fluid pH

Calculus formation
Periodontal
disease

Mineralised plaque
a deposit of
calcium phosphate salts
on the tooth surface

Classification
supra-gingival calculus
sub-gingival calculus

Composition
Inorganic compound 80-90%
Organic compound:
Proteins,Lipids,Carbohydrates Oxalic
acid, Porphyrins
Source of compounds
Saliva,
Gingival crevicular fluid
Food, Bacteria

Hydroxyapatite
Ca10(PO4)6(OH)2
Whitlockite or tricalcium phosphate
(-TCP) Ca3(PO4)2
Octacalcium phosphate (OCP)
Ca8H2(PO4)6.5H2O
Brushite or
dicalcium phosphate dihydrate (DCPD)
CaHPO4.2H2O

carbonate apatite
(Ca,M)10(CO3,HPO4,PO4)6(OH,X)2
M = metal ion eg. Mg, X = Cl, F

Amorphous calcium phosphate (ACP)

Calcium phosphate phases

more soluble phases and/or
larger crystals
lower attachment to apatite crystals
on tooth surfaces

easier to remove

Proteins
osteopontin (calcium-binding
phosphorylated glycoprotein
produced by osteoblast)
calprotectin (calcium-binding protein
produced by leukocytes, macrophage and
epithelial cell)

Lipids
%
Neutral lipid

61.8

(free fatty acid)

Glycolipid

28.0

(simple glycosphingolipids 17.2

neutral and sulfated glyceroglucolipids 82.8)

Phospholipid

10.2

(phosphatidylethanolamine 34.2%, diphosphatidylglycerol

25.5%,
phosphatidylinositol 2.3%
phosphatidylserine 1.7%)

Formation
Supra-gingival calculus

Sub-gingival calculus

Supra-gingival plaque

Sub-gingival plaque

Interaction

Interaction

Saliva

Gingival crevicular fluid

3 Factors in Calculus Formation

1. Physicochemical change
2. Microorganism activity
3. Modification of inhibitors and
inhibitor-destruction mechanism

1. Physicochemical change
pH
Calcium and phosphate conc.
pH

Ca, PO4

Ion product (Ip)

Solubility product (Kp)
Saturation degree (SD)
Ip >> Kp
precipitation
Ip << Kp
solubilization

2. Microorganism activity
pH
Microbial products eg.

Lactic acid
Urea
Calcium

Structure

Role
Periodontal
disease

Prevention
Triclosan with Polyvinylmethyl ether
(PVM)/ Maleic acid (MA) copolymer

Pyrophosphate and PVM/MA

copolymer
Zinc ion

Triclosan
Broad spectrum antibacterial agent
both Gram +ve and -ve
Bactericidal effect: target at cell
membrane
Bacteriostatic effect: inhibit uptake of
essential amino acids

Anti inflammation
Inhibition: cyclooxygenase,
lipooxygenase
PVM/MA copolymer
Delivery system
Weak inhibitor of crystal growth

Pyrophosphate
Inhibit crystal growth
By binding to HA surface
Prevent binding of phosphate to HA surface

Zinc ion
Reversible inhibit crystal growth

An inverse relationship between

calculus and caries
Caries: demineralization
Calculus: mineralization

Relationship between caries and calculus prevalence

Trial

Calculus-free
DMSF

Calculus-prone
DMSF

Caries diff*
(%)

Bristol

8.85

7.11

19.7

Berkshire

9.22

8.09

12.3

Munster

12.31

10.48

14.9

Somerset

6.62

5.60

15.4

Villefranche

5.62

5.53

1.6

Lanarkshire

11.08

8.40

24.2

*DMSF (CF) DMSF (CP) / DMSF (CF)