Food Biochemistry and Food Processing

9 Protein Cross-linking in Food 225

processing regime. Those that react do so with dif-
fering degrees of reactivity under various condi-
tions.

DISULFIDECROSS-LINKS

Disulfide bonds are the most common and well-
characterized type of covalent cross-link in proteins
in biology. They are formed by the oxidative coupling
of two cysteine residues that are close in space within
a protein. A suitable oxidant accepts the hydrogen
atoms from the thiol groups of the cysteine residues,
producing disulfide cross-links. The ability of pro-
teins to form intermolecular disulfide bonds during
heat treatment is considered to be vital for the gelling
of some food proteins, including milk proteins, suri-
mi, soybeans, eggs, meat, and some vegetable pro-
teins (Zayas 1997). Gels are formed through the
cross-linking of protein molecules, generating a
three-dimensional solid-like network, which pro-
vides food with desirable texture (Dickinson 1997).
Disulfide bonds are thought to confer an element
of thermal stability to proteins and are invoked, for
example, to explain the heat stability of hen egg
white lysozyme, which has four intramolecular di-
sulfide cross-links in its native conformation (Ma-
saki et al. 2001). This heat stability influences many
of the properties of egg white observed during cook-
ing. Similarly, the heat treatment of milk promotes
the controlled interaction of denatured
-lactoglob-
ulin with -casein, through the formation of a disul-
fide bond. This increases the heat stability of milk
and milk products, preventing precipitation of
-
lactoglobulin (Singh 1991). Disulfide bonds are also
important in the formation of dough. Disulfide inter-
change reactions during the mixing of flour and
water result in the production of a protein network
with the viscoelastic properties required for bread
making (Lindsay and Skerritt 1999). The textural
changes that occur in meat during cooking have also
been attributed to the formation of intermolecular
disulfide bonds (Singh 1991).

CROSS-LINKSDERIVED FROM
DEHYDROPROTEIN

Alkali treatment is used in food processing for a
number of reasons, such as the removal of toxic con-
stituents, and the solubilization of proteins for the
preparation of texturized products. However, alkali
treatment can also cause reactions that are undesir-

able in foods, and its safety has come into question

(Friedman 1999a,c; Shih 1992; Savoie 1991). Ex-

posure to alkaline conditions, particularly when cou-

pled to thermal processing, induces racemization of

amino acid residues and the formation of covalent

cross-links, such as dehydroalanine, lysinoalanine,

and lanthionine (Friedman 1999a,b,c). Dehydroal-

anine is formed from the base-catalyzed elimination

of persulfide from an existing disulfide cross-link.

The formation of lysinoalanine and lanthionone

cross-links occurs through
-elimination of cysteine

and phosphoserine protein residues, thereby yielding

dehydroprotein residues. Dehydroprotein is very re-

active with various nucleophilic groups including the

-amino group of lysine residues and the sulfhydryl

group of cysteine. In severely heat- or alkali-treated

proteins, imidazole, indole, and guanidino groups of

other amino acid residues may also react (Singh

1991). The resulting intra- and intermolecular cross-

links are stable, but food proteins that have been

extensively treated with alkali are not readily digest-

ed, reducing their nutritional value. Mutagenic prod-

ucts may also be formed (Friedman 1999a,c).

CROSS-LINKSDERIVED FROMTYROSINE

Various cross-links formed between two or three

tyrosine residues have been found in native proteins

and glycoproteins, for example in plant cell walls

(Singh 1991). Dityrosine cross-links have recently

been identified in wheat, and are proposed to play a

role in formation of the cross-linked protein network

in gluten (Tilley 2001). They have also been formed

indirectly, by treating proteins with hydrogen perox-

ide or peroxidase (Singh 1991), and are implicated

in the formation of caseinate films by gamma irradi-

ation (Mezgheni et al. 1998). Polyphenol oxidase

can also lead indirectly to protein cross-linking, due

to reaction of cysteine, tyrosine, or lysine with reac-

tive benzoquinone intermediates generated from the

oxidation of phenolic substrates (Matheis and

Whitaker 1987, Feeney and Whitaker 1988). Such

plant phenolics have been used to prepare cross-

linked gelatin gels to develop novel food ingredients

(Strauss and Gibson 2004).

CROSS-LINKSDERIVED FROM THEMAILLARD

REACTION

The Maillard reaction is a complex cascade of chem-

ical reactions, initiated by the deceptively simple