BLBS102-c14 BLBS102-Simpson March 21, 2012 13:17 Trim: 276mm X 219mm Printer Name: Yet to Come
264 Part 2: Biotechnology and EnzymologyPROTEASES
Proteases play an important role in the growth and survival of all
living organisms. Protease catalyzes the hydrolysis of peptide
bonds in polypeptides and protein molecules (Garcia-Carreno
and Hernandez-Cortes 2000). On the basis of their in vitro prop-
erties, proteases have been classified in a number of ways such as
the pH range over which they are active (acid, neutral, or alkaline
proteases), their ability to hydrolyze specific proteins, and their
mechanism of catalysis. Proteases may be classified based on
their similarities to well-characterized proteases such as trypsin-
like, chymotrypsin-like, chymosin-like or cathepsin-like, and so
on (Haard 1990). In addition, proteases are classified according
to their catalytic action (endopeptidase or exopeptidase) and the
nature of the catalytic site. In Enzyme Commission (EC) system
for enzyme nomenclature, all proteases (peptide hydrolases) be-
long to subclass 3.4, which is further divided into 3.4.11–19,
the exopeptidases and 3.4.21–24, the endopeptidases or pro-
teinases (Nissen 1993). Endopeptidases cleave the polypeptide
chain at particularly susceptible peptide bonds distributed along
the chain, whereas exopeptidases hydrolyze one amino acid unit
sequentially from theN-terminus (aminopeptidases) or fromC-
terminus (carboxypeptidases) (Fig. 14.1). Exopeptidases, espe-
cially aminopeptidases, are ubiquitous, but less readily available
as commercial products, since many of them are intracellular or
membrane bound (Simpson 2000). For endopeptidases or pro-
teinases, the four major classes of endopeptidases can be dis-
tinguished according to the chemical group of their active site,
including serine proteinases (EC 3.4.21), thiol or cysteine pro-
teinases (EC 3.4.22), acid or aspartic proteinases (EC 3.4.23),
and metalloproteinases (EC 3.4.24) (Simpson 2000). The en-
zymes in the different classes are differentiated by various cri-
teria, such as the nature of the groups in their catalytic sites,
their substrate specificity, and their response to inhibitors or by
their activity/stability under acid or alkaline conditions (Nissen
1993).
For marine animals, proteases are mainly produced by the
digestive glands. Like the proteases from plants, animals, and
microorganisms, digestive proteases from marine animals are
polyfunctional enzymes catalyzing the hydrolytic degradation
of proteins (Garcia-Carreno and Hernandez-Cortes 2000). Forsome fish species, proteases are present at high levels in the mus-
cle (Kolodziejska and Sikorski 1996), and are associated with
the induced changes of proteins during postmortem storage or
processing. Marine animals have adapted to different environ-
mental conditions, and these adaptations, together with inter-
and intraspecies genetic variations, are associated with certain
unique properties of their proteinases, compared with their coun-
terpart from land animals, plants, and microorganisms (Simpson
2000). Some of these distinctive properties include higher cat-
alytic efficiency at low temperature and lower thermal stability
(Klomklao et al. 2009a).Digestive Proteases from Marine AnimalsDigestive proteases have been studied in several species of
fish and decapods (De-Vecchi and Coppes 1996). Proteases
found in the digestive organs of fish include pepsin, gastricsin,
trypsin, chymotrypsin, collagenase, elastase, carboxypeptidase,
and carboxyl esterase (Simpson 2000). Pepsin, chymotrypsin,
and trypsin are three main groups of proteases found in fish
viscera. Pepsin is localized in fish stomach (Klomklao et al.
2007a), while chymotrypsin and trypsin are concentrated in tis-
sues such as the pancreas, pyloric ceca, and intestine (Klomklao
et al. 2004). The distribution and properties of protease vary
depending on species and organs (Table 14.1). According to
the International Union of Applied Biochemists classification,
digestive proteases from fish and aquatic invertebrates may be
classified into four major groups such as acidic/aspartic protease,
serine protease, thiol or cysteine protease, and metalloprotease
(Simpson 2000).Acid/Aspartic ProteasesThe acid or aspartic proteases are a group of endopeptidases
characterized by high activity and stability at acidic pH. They
are referred to as “aspartic” proteases (or carboxyl proteases)
because their catalytic sites are composed of the carboxyl group
of two aspartic acid residues (Whitaker 1994). On the basis
of the EC system, all the acid/aspartic proteases from marine
animals have the first three digits in common as EC 3.4.23.
Three common aspartic proteases that have been isolated andExopeptidaseEndopeptidase +
Figure 14.1.Cleavage of proteins by endopeptidases and exopeptidases.