Food Biochemistry and Food Processing (2 edition)

BLBS102-c09 BLBS102-Simpson March 21, 2012 11:15 Trim: 276mm X 219mm Printer Name: Yet to Come

9 Enzymes in Food Processing 189

modification of gluten. The enzymes are able to do this by in-

creasing the viscoelastic, gas-retaining, and thermosetting prop-

erties of the dough. The action of these enzymes results in bene-

fits such as shorter time of mixing, better flow, improved shaping,

and increased gas retention of the flour or dough (Mathewson

1998).

Metalloproteases

The term metalloproteases is used to encompass those proteases

that have metal ions in their active sites. These enzymes typically

require an essential metal ion (e.g., Ni^2 +,Mg^2 +,Mn^2 +,Ca^2 +,

Zn^2 +, and Co^2 +) for functional activity. Urease is an example

of the nickel (Ni^2 +)-containing metalloenzymes. It catalyzes the

breakdown of urea into carbon dioxide (CO 2 ) and ammonia

(NH 3 ). The urease-catalyzed reaction occurs as follows:

(NH 2 ) 2 CO+H 2 O→CO 2 +2NH 3

The NH 3 thus produced functions to neutralize gastric acid

in the stomach. Urease is found in bacteria, yeast, and several

higher plants. The major sources of urease include the Jack bean

(Canavalia ensiformis) plant and the bacteriaB. pasteurii.The

enzyme is also found in large quantities in soybeans and other

plant seeds, in certain animal tissues (e.g., liver, blood, and mus-

cle), in intestinal bacteria (e.g.,Eubacterium aerofaciens,E.

lentum,andPeptostreptococcus productus), and in yeasts (e.g.,

Candida curvata, C. humicola, C. bogoriensis,C. diffluens,and

RhodotorulaandCryptococcusspecies), as well as the blue-

green algae (Anabaena doliolumandAnacystis nidulans). Ex-

amples of the metalloproteases that are Mg^2 +dependent include

inorganic diphosphatase that catalyzes the cleavage of diphos-

phate into inorganic phosphates, that is,

Diphosphate+H 2 O⇔2 Phosphate

and phosphatidylinositol triphosphate 3-phosphatase that

catalyzes the hydrolysis of phosphatidylinositol 3,4,5-

trisphosphate, that is,

Phosphatidylinositol 3, 4 ,5-trisphosphate+H 2 O

⇔Phosphatidylinositol 4,5-bisphosphate+Phosphate

An example of metalloproteases containing manganese

(Mn^2 +) metal ions is phosphoadenylylsulfatase, which partici-

pates in sulfur metabolism and catalyzes the hydrolytic cleavage

of 3′-phosphoadenylyl sulfate into adenosine 3′,5′-bisphosphate

and sulfate:

3 ′-Phosphoadenylyl sulfate+H 2 O

⇔Adenosine 3′, 5 ′-Bisphosphate+Sulfate

A well-known calcium (Ca^2 +)-containing metalloprotease is

lactonase, which hydrolyzes lactones into their corresponding

hydroacids, that is,

Lactone(s)+H 2 O⇔4-Hydroxyacid(s)

The enzyme lactonase also participates in the metabolism of

galactose and ascorbic acid.

Other metalloproteases contain zinc (Zn^2 +) metal ions,

e.g., glutamate carboxypeptidase and glutamyl aminopepti-

dase. Glutamate carboxypeptidase catalyzes the hydrolysis

ofN-acetylaspartylglutamic acid into glutamic acid andN-

acetylaspartate as shown in the following reaction:

N-Acetylaspartylglutamate+H 2 O

⇔Glutamic acid+N-Acetylaspartate

while glutamyl aminopeptidase (also known as aminopeptidase

A) catalyzes the hydrolysis of glutamic and aspartic acid residues

from the N-termini of polypeptides, e.g.,

Aspartic acid-polypeptide+H 2 O

⇔Aspartic acid+Polypeptide

Glutamyl aminopeptidase is important because of its capacity

to degrade angiotensin II into angiotensin III, which promote

vasoconstriction and high blood pressure. Both glutamate car-

boxypeptidase and glutamyl aminopeptidase are present in mem-

branes. Another important Zn^2 +-containing metalloprotease is

theinsulin-degrading enzymethat is found in humans and de-

grades short chain polypeptides such as theβ-chain of insulin,

glucagon, transforming growth factor, and endorphins. Collage-

nases are also Zn^2 +dependent and they act to break down the

peptide bonds in collagen. Cobalt (Co^2 +)-containing metallo-

proteases include methylmalonyl coenzyme A mutase that con-

verts methyl malonyl-CoA to succinyl-CoA, which is involved

in the extraction of energy from proteins and fats; other Co^2 +-

containing metalloenzymes are methionine aminopeptidase and

nitrile hydratase (Kobayashi and Shimizu 1999).

Sources of metalloproteases include those derived from an-

imals, like carboxypeptidases A and B, collagenases, gelati-

nases, and procollagen proteinase; microbial ones such as

thermolysin, lysostaphin (fromStaphylococcussp), acidolysin

(fromClostridium acetobutylicum), and neutral metallopro-

teases (fromBacillussp); as well as plant types like the zinc-

dependent endoproteases in the seeds of grass pea (Lathyrus

sativusL.). Agents or substances that are capable of bind-

ing or interacting with the catalytic metal ions tend be po-

tent inhibitors of this group of enzymes. Examples of these

inhibitors include phosphoramidon, ethylenediaminetetraacetic

acid (EDTA), methylamine, vitamin B-12, dimercaprol, por-

phine, phosphonates, and 1,10-phenanthroline.

Common commercially available metalloenzymes are Neu-

trase (Novo Nordisk 1993) and Thermoase (Amano Enzymes)

and some of the industrial applications of metalloenzymes in-

clude the synthesis of peptides for use as low calorie sweetener

and drugs for the food and pharmaceutical industries, respec-

tively. Thermolysin and vimelysin (fromVibriosp) are both

used for the synthesis of aspartame (Murakami et al. 1998); in

this regard, vimelysin is superior to thermolysin for synthesis

of the dipeptide at low temperatures. The metalloprotease ther-

molysin is also used in combination with other proteases (pro-

tease cocktails) for the production of food protein hydrolysates

and flavor-enhancing peptides and for accelerating the ripening

of dry sausages (Fernandez et al. 2000). They are also used for

the hydrolysis of slaughterhouse waste, and in the brewing in-

dustry to facilitate filtration of beer and production of low calorie

beers.