Food Biochemistry and Food Processing (2 edition)

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304 Part 3: Meat, Poultry and Seafoods

are located close enough in the real meat product for an effective

interaction, and (4) the enzyme exhibits enough stability during

processing for the changes to be developed.

DESCRIPTION OF THE MUSCLE

ENZYMES

There are a wide variety of enzymes in the muscle. Most of

them have an important role in the in vivo muscle functions,

but they also serve an important role in biochemical changes

such as the proteolysis and lipolysis that occur in postmortem

meat, and during further processing of meat. Some enzymes are

located in the lysosomes, while others are free in the cytosol or

linked to membranes. The muscle enzymes with most important

roles during meat processing are grouped by families and are

described in the succeeding sections.

Muscle Proteases

Proteases are characterized by their ability to degrade proteins,

and they receive different names depending on respective mode

of action (see Fig. 16.1). They are endoproteases or proteinases,

when they are able to hydrolyze internal peptide bonds, but they

are exopeptidases, when they hydrolyze external peptide bonds,

either at the amino termini or the carboxy termini.

Neutral Proteinases: Calpains

Calpains are cysteine endopeptidases consisting of heterodimers

of 110 kDa, composed of an 80 kDa catalytic subunit and a

30 kDa subunit of unknown function. They are located in the

cytosol, around the Z-line area. Calpains have received differ-

ent names in the scientific literature, such as calcium-activated

neutral proteinase, calcium-dependent protease, and calcium-

O O O

C

C

HN

C

C

HN

C

C

HN

O

OH

O

C

C

HN

C

C

H 3 N

n

O O

Endopeptidase

Dipeptidylpeptidase

HN C

C

C

O

HN

C

C

HN

O

OH

O

C

C

HN

C

C

H 3 N

n

C HN C C

O

OH

O

C C

n

Aminopeptidase Carboxypeptidase

Figure 16.1.Mode of action of the different types of muscle

proteases.

activated factor. Calpain I is also calledμ-calpain because it

needs micromolar amounts (50–70μM) of Ca^2 +for activation.

Similarly, calpain II is calledm-calpain because it requires mil-

limolar amounts (1–5 mM) of Ca^2 +. Both calpains show max-

imal activity around pH 7.5. Calpain activity decreases very

quickly when pH decreases to 6.0, or even reaches ineffective

activity at pH 5.5 (Etherington 1984). Calpains have shown

good ability to degrade important myofibrillar proteins, such as

titin, nebulin, troponins T and I, tropomyosin, C-protein, filamin,

desmin, and vinculin, which are responsible for the fiber struc-

ture. On the other hand, they are not active against myosin, actin,

α-actinin and troponin C (Goll et al. 1983, Koohmaraie 1994).

The stability of calpain I in postmortem muscle is very poor

because it is readily autolyzed, especially at high temperatures,

in the presence of the released Ca^2 +(Koohmaraie 1994). Calpain

II appears as more stable, just 2–3 weeks before losing its activity

(Koohmaraie et al. 1987). In view of this rather poor stability,

the importance of calpains should be restricted to short-term

processes. A minor contribution, just at the beginning, has been

observed in long processes such as dry curing of hams (Rosell

and Toldr ́a 1996) or in fermented meats where the acid pH values

makes calpain activity rather unlikely (Toldra et al. 2001). ́

Calpastatin is a polypeptide (between 50 and 172 kDa) acting

as an endogenous reversible and competitive inhibitor of calpain

in the living muscle. In postmortem muscle, calpastatin regulates

the activity of calpains, through a calcium-dependent interaction,

although only for a few days, because it is destroyed by autolysis

(Koohmaraie et al. 1987). The levels of calpastatin vary with

animal species, and pork muscle has the lowest level (Valin and

Ouali 1992).

Lysosomal Proteinases: Cathepsins

There are several acid proteinases in the lysosomes that degrade

proteins in a nonselective way. The most important are cathep-

sins B, H, and L, which are cysteine proteinases, and cathepsin

D, which is an aspartate proteinase. The optimal pH for activity

is slightly acid (pH around 6.0) for cathepsins B and L, acid (pH

around 4.5) for cathepsin D, and neutral (pH 6.8) for cathepsin

H(Toldra et al. 1992). Cathepsins require a reducing environ- ́

ment such as that found in postmortem muscle to express their

optimal activity (Etherington 1987). All of them are of small

size, within the range 20–40 kDa, and are thus able to penetrate

into the myofibrillar structure. Cathepsins have shown a good

ability to degrade different myofibrillar proteins. Cathepsins D

and L are very active against myosin heavy chain, titin, M and C

proteins, tropomyosin, and troponins T and I (Matsukura et al.

1981, Zeece and Katoh 1989). Cathepsin L is extremely active

in degrading both titin and nebulin. Cathepsin B is able to de-

grade myosin heavy chain and actin (Schwartz and Bird 1977).

Cathepsin H exhibits both endo- and aminopeptidase activity,

and this is the reason for its classification as an aminoendopep-

tidase (Okitani et al. 1981). In the muscle, there are endogenous

inhibitors against cysteine peptidases. These inhibitory com-

pounds, known as cystatins, are able to inhibit cathepsins B, H,

and L. Cystatin C and chicken cystatin are the most well-known

cystatins.