Papain like proteases : Applications of their inhibitors

Text-only Preview

African Journal of Biotechnology Vol. 6 (9), pp. 1077-1086, 2 May 2007
Available online at
ISSN 1684–5315 © 2007 Academic Journals

Papain-like proteases: Applications of their inhibitors

Vikash K. Dubey1*, Monu Pande2, Bishal Kumar Singh1 and Medicherla V. Jagannadham2

1Department of Biotechnology, Indian Institute of Technology, Guwahati- 781039, Assam, India.
2Molecular Biology Unit, Institute of Medical Sciences, Banaras Hindu University, Varanasi-221005, India.

Accepted 5 February, 2007

Proteases are one of the most important classes of enzyme and expressed throughout the animal and
plant kingdoms as well as in viruses and bacteria. The protease family has drawn special attention for
drug target for cure of several diseases such as osteoporosis, arthritis and cancer. Many proteases
from various sources are being studied extensively with respect to activity, inhibition and structure. In
this review, we hope to bring together the information available about the proteases with particular
emphasis on papain-like plant cysteine proteases. Besides, protease inhibitors and their potential
utilities are also discussed.

Key words:
Proteases, plant latex, reaction mechanism and protease inhibitors.


Proteolytic enzymes are of widespread interest because
shows evidence of their evolutionary relationship by their
of there industrial application and because they have
similar tertiary structures, by the order of catalytic resid-
been implicated in the design and synthesis of thera-
ues in their sequences, or by common sequence motifs
peutic agents (Neurath, 1989). With the advent of mole-
around the catalytic residues. The proteases have been
cular biology, proteolytic enzymes have become a fertile
organized in to evolutionary families and clans by
and exciting field of basic as wel as applied research.
Rawlings and Barrett (1993, 1994), which led to develop-
Identification of novel genes encoding proteases has
ment of MEROPS database of proteases. MEROPS
considerably increased our knowledge of proteases and
database ( includes listing of
provided fresh imputes. The proteases have different cel-
al peptidase sequences from different families and clans.
lular distribution and intracel ular localization which may
Each new updates adds new members and families.
contribute to defining specific functional roles for some of
Some representative family and clans of cysteine, serine
these proteases.
and threonine proteases are listed in Table 1. The related
Proteases have been divided into six mechanistic
families are grouped into clans, which contains al the
classes by the International Union of Biochemistry. These
peptidase that arose from a single evolutionary origin.
include the cysteine, serine, aspartic, metal oprotease,
The designation of family fol ows the catalytic type, serine
threonine and unknown type (Enzyme nomenclature,
(S), cysteine (C), or threonine (T). However, some of the
1992). The threonine protease is the most recently disco-
clans are mixed type and contains families with two
vered (Seemul er et al., 1995). Each class has a charac-
catalytic types or more catalytic types and designated
teristic set of functional amino acid residues arranged in a
with the letter “P”. The cysteine protease family compri-
particular configuration to form the active site. The diffe-
ses six major families: the papain family, calpains, clostri-
rent proteases class includes distinct families and the
pains, streptococcal cysteine proteases, viral cysteine
members from different family differ from each other in
proteases and most recently established, caspases (also
amino acid sequence despite a common active site geo-
cal ed apopains). Overal , twenty families of cysteine
metry and enzymatic mechanism. Family of peptidases
peptidases have been recognized (Rawlings and Barrett,

1994). The order of cysteine and histidine residues

(Cys/His or His/Cys) in the linear sequence differs

between families. The families C1, C2 and C10 can be
*Corresponding author. E-mail: vdubey@i Tel: +91-
described as papain-like, C3, C4, C5, C6, C7, C8, C9,
361-2582203. Fax: +91-361-2582249.
C16, C18 and C21 are represented in viruses while C11,

1078 Afr. J. Biotechnol.

C15 and C20 are from bacterial source.
and viral cysteine proteases with the trypsin-fold are

classified as ?-proteins. The proteasome subunits are ? +

? proteins composed mainly of antiparal el ?-sheets with
segregated ? and ? regions. The group of cysteine

proteases with papain, cruzain, and cathepsin also has
Hydrolysis of a peptide bond is an energetical y favorable
this structure. The subtilisins and caspases are members
reaction, but extremely slow (Wolfenden and Snider,
of the ?/? group of proteins with paral el ?sheets (?-?-?
2001). The active site residues of serine, cysteine, and
units). Al known cysteine proteases can be grouped in at
threonine proteases are shown in Figure 1A. The active
least 30 protein families. Each family contains proteins
site residues in al class of proteases have many
with similar amino acid sequences and evolutionarily
mechanistic features in common. Each enzyme has an
conserved sequence motifs, which reflects the family
active site nucleophile and a basic residue, which can
members' similar 3D structures.
also function as a general acid in the catalytic mecha-
Three-dimensional structure has been elucidated for
nism. The transition states for serine, cysteine, and threo-
papain, a representative member of papain-like cysteine
nine proteases al involve formation of a tetrahedral
proteases (Drenth et al., 1971; Kamphuis et al., 1984),
intermediate shown in Figure 1B. The oxyanion of the
as wel as other members like actinidin (Baker, 1980),
tetrahedral intermediate is frequently stabilized by inter-
calotropin (Heinemann et al., 1982), cathepsin B (Musil et
action with several hydrogen bond donors, which is
al., 1991), caricain (Pickersgil et al., 1991), glycyl
commonly referred to as the oxyanion hole. The oxyanion
endopeptidase or papaya proteinase IV (O’Hara et al.,
hole of serine proteases is usual y quite rigid and involves
1995), chymopapain (Maes et al., 1996) and cruzain
backbone peptide bond NH groups as hydrogen bond
(McGrath et al., 1995) and al show bilobed molecules in
donors. Interaction with the oxyanion hole is usual y
which catalytic site is located in a cleft between the lobes.
essential for effective substrate hydrolysis. With cysteine
Al papain-like cysteine proteases share similar seque-
proteases, the oxyanion hole does not seem to be as
nces (Kamphuis et al., 1985; Kirschke et al., 1995; Berti
essential and is much more flexible at least in the case of
and Storer, 1995) and have similar 3-dimensional struc-
the papain family.
tures. The structural data provides a strong evidence that
Cysteine peptidases of the papain family catalyze the
al arose from a common ancestor. Al known papain-like
hydrolysis of peptide, amide, ester, thiol ester and thiono
cysteine proteases, irrespective or origin, except cathe-
ester bonds (Brocklehurst et al., 1987). The basic fea-
psin C, are monomers whose structure consists of two
tures of the mechanism include the formation of a cova-
domains (referred to as the R- and L- domains) according
lent intermediate, the acyl-enzyme, resulting from nucleo-
to their right and left position in the standard view. The
philic attack of the active site thiol group on the carbonyl
domains fold together in the form of a closed book. The
carbon of the scissile amide or ester bond of the bound
interactions between the domains have hydrophobic as
substrate. The first step in the reaction pathway corres-
wel as hydrophilic character and are specific for a
ponds to the association (or noncovalent binding) of the
particular enzyme. The structure of papain has been
free enzyme and substrate to form the Michaelis comp-
extensively studied. The two catalytic residues that is,
lex. Acylation of the enzyme, with formation and release
Cys 25 and His 159 in papain each from N- and C-
of a first product fol ow this step from the enzyme, the
terminal domains respectively, are present in a ‘V’ like
amine R?NH2. In the fol owing step, the acyl-enzyme
shaped active site cleft situated on the top of the enzyme
reacts with a water molecule to form the second product
structure. Recently structure of two new of papain-like
(deacylation step). Release of this product results in the
cysteine proteases, ervatamin B and ervatamin C,
regeneration of the free enzyme.
purified in our laboratory, have been reported (Biswas et

al., 2003; Thakurta et al., 2004), which also shows strong

structural similarly with papain (Figure 2).
Papain has a large binding site and there are a number

of interactions that exist between the enzyme and the
The structural determination (x-ray of NMR) of proteases
substrate over an extended region. Coupling of these
is lagging considerably behind the sequence determina-
substrate binding interactions to the hydrolytic process
tion. MEROPS database indicates that the structures of
occurring at the active site is an important aspect of
majority of proteases are not yet available and opens a
catalysis. Schechter and Berger (1967) proposed that the
wide area of studies. However, the available structures
active site of papain contained seven subsites each
show high degree of variability. The proteases seem to
capable of accommodating a single amino acid residue of
be distributed into al of the major structural classes of
a peptide substrate. The subsites are located on both
proteins [?-proteins, ?-proteins, ?- and ?- proteins (?/? or
sides of the catalytic site, four on the N-terminal side and
? + ?), multidomain proteins, membrane and cel surface
three on the C-terminal side. The amino acid residues on
proteins, and smal proteins]. Prokaryotic and eukaryotic
the amino-terminal side of the scissile bond are num-
trypsin-like serine proteases, some viral serine proteases,
bered P1, P2, P3,… counting outwards; the residues on

Dubey et al. 1079

Table 1. Representative families and clans of cysteine, serine and threonine proteases.

C1, C2
papain, cathepsins B, K,L,S,H
C11, C13, C14, C25, C50
legumain, caspases. gingipain, separase
C3 (viral), C30 (viral)
picornain 3C, SARS virus 3C-like endopeptidase
chymotripsin, trypsin, elastase, cathepsin G
archaean proteasome
S9, S10
Prolyl oligopeptidase, carboxypeptidase Y
S11, S12
D-Ala-D-Ala carboxypeptidases A and B

Rawlings and Barrett (1993, 1994).


Figure 1. (A) Active site residue (B) transition states of protease hydrolysis of serine, cysteine,
and threonine proteases. In serine proteases, three residues form the catalytic triad are essential
in the catalytic process, that is, His 57, Asp 102 and Ser 195 (chymotrypsinogen numbering). In
cysteine proteases, catalysis proceeds through the formation of a covalent intermediate and
involves a cysteine and a histidine residue. While active site of threonine proteases comprises of
threonine, methionine and backbone amide.

the carboxy-terminal side of the scissile bond are
papain one of the subsites, S2, specifical y interacts with
numbered P11, P21, P31,... The subsites on the protease
a phenylalanine side chain of peptides.
are termed S3, S2, S1, S11, S21, S31... to complement
Thus, al members of the papain family inspite of their
the substrate residues that interact with the enzyme. In
dissimilar origin, show considerable similarity in terms of

1080 Afr. J. Biotechnol.

Figure 2.
Ribbon diagram of (A) Papain (PDB accession: 9PAP), (B) Ervatamin B (PDB
accession: 1IWD) and (C) Ervatamin C (PDB accession: 1O0E) showing catalytic residues i.e.,
Cys and His between cleft of two domain. Al papain-like cysteine proteases have catalytic residue
between domain cleft.

activity, pH optima, molecular mass, catalytic mechanism
to the papain family sharing similar protein structure and
and the peptide regions near the active site cysteines are
mechanism of action. However, slight structural differen-
quite similar. However, despite the similarities, it has
ces make these enzymes distinct with respect to their
been suggested that al the cysteine proteases may have
substrate specificity and regulation. Cathepsins are syn-
arisen by convergent evolution (Lowe, 1976). More struc-
thesized as 30 - 50 kDa precursor proteins, which are
tural information from sulfhydryl enzymes of more closely
glycosylated and phosphorylated in the Golgi apparatus.
related genera of plants and microbes is needed to
They are processed in the lysosomes to their active
conclude that whether or not any evolutionary relation-
forms by one or more proteolytic cleavage. The optimum
ship exists. Structural determination of several proteases
activity of cathepsins is pH 5.0 - 6.5, although they can
purified in our laboratory may provide additional insights
hydrolyze large substrates also at neutral pH. The pH
(Dubey and Jagannadham, 2003a; Patel and Jaganna-
dependent activity of cathepsins is rather complex and
dham, 2003; Nal amsetty et al., 2003).
depends not only on the microenvironment and the

nature of the conformation of the substrate, but also on

the presence or absence of stabilizing factors (Keppler
and Sloane, 1996).
Most of these papain-like enzymes are relatively smal

proteins with Mr values in the range 20 - 35 kDa (Bro-
Cysteine proteases of the papain super family are widely
cklehurst et al., 1987; Polgar et al., 1989; Rawlings and
distributed in nature. They can be found in both prokary-
Barrett, 1994; Berti and Storer, 1995). However, cathe-
otes and eukaryotes e.g. bacteria, parasites, plant,
psin C is an oligomeric enzyme with Mr ~ 200 kDa
invertebrates and vertebrates (Berti and Storer, 1995).
(Metrione et al., 1970). Al cysteine proteases except
Papain-like cysteine proteases are the most abundant
cathepsin C are endopeptidases (Kirschke et al., 1995).
among the cysteine proteases. The papain family
Cathepsin B is a dipeptidyl carboxypeptidase (Aronson
contains peptidases with a wide variety of activities,
and Barrett, 1978), cathepsin H is an aminopeptidase
including endopeptidases with broad specificity (such as
(Koga et al., 1992) and cathepsin C is a dipeptidyl amino-
papain), endopeptidases with very narrow specificity
peptidase but at higher pH exhibits dipeptidyl transferse
(such as glycyl endopeptidases), aminopeptidases, a
activity (Kirschke et al., 1995).
dipeptidyl-peptidase, and peptidases with both endopep-
Disturbance of the normal balance of enzymatic activity
tidase and exopeptidase activities (such as cathepsins B
of lysosomal cysteine proteases may lead to pathological
and H). There are also family members that show no
conditions, and these proteases have been found to be
catalytic activity. Enzymes of papain family are found in a
involved in many such cases. The participation of these
wide variety of life forms: baculovirus (Rawlings et al.,
enzymes in various diseases seems to be restricted to
1992), eubacteria like Porphyromonas and Lactococcus,
their proteolytic function outside the lysosomes, after
yeast (Enenkel and Wolf, 1993), and probably al
secretion from lysosomes or after translocation into
protozoa, plants, and animals.
different intracel ular granules. The resulting uncontrol ed
The family consists of papain and related plant prote-
proteolysis is a result of an imbalance between catalyti-
ases such as chymopapain, caricain, bromelain, actinidin,
cal y active proteases and their natural inhibitors, and can
ficin, aleurain, etc. Lysosomal cysteine proteases, also
be observed in e.g. inflammation and tumor growth,
known as cysteine cathepsins (Cats), include Cat B, Cat
although these processes are very complex.
H, Cat S, Cat K, Cat O/2, Cat F, Cat W and Cat U
Cysteine proteases of the papain family have been
(Chapman et al., 1997; Turk et al., 1997) and also belong
reported in bacteria as wel . Proteolytic enzymes produ-

Dubey et al. 1081

ced by Porphyromonas gingivalis are important virulence
melain (Takahasi et al., 1973), papain (Kimmel and
factors of this periodontopathogen. In the periodontal
Smith, 1954), and ficin (Englund et al., 1968) and have
disease proteolytic enzymes are produced in large
used extensively in food and medicine industry. Besides,
quantities. It has been shown that these proteases can
some of these proteases have also been used as model
directly or indirectly degrade constituents of the period-
systems for studies on structure-function relationships
ontal tissues, destroy host defense elements, dysregulate
and protein folding problems (Kundu et al., 1999; Edwin
coagulation and complement kal ikerinkinin cascades.
and Jagannadham, 1998, 2000; Dubey and Jaganna-
Recently, proteases belonging to two catalytic classes
dham 2003b). Proteolytic enzymes from plant sources
and produced by P. gingivalis have been identified. One
have received special attention in the pharmaceutical
enzyme is described as an Arg-X specific proteinase
industry and biotechnology due to their property of being
(Chen et al., 1992) and another is Lys-X specific (Pike et
active over wide ranges of temperature and pH. Al the
al., 1994). Since the first purified enzyme shared some
plant cysteine proteases exhibit pH optima in the region
properties with clostripain, it was named as a gingipain.
5.0 - 8.0, and almost al have a molecular mass in the
Fol owing the recommendations by the IUB, these protea-
range 25 - 30 kDa except a few in the range of 50 - 75
ses are referred to as gingipain-R and gingipain-K to
kDa. It is probable that al such sulfhydryl endopeptidases
account for their unique specificity.
employ similar catalytic mechanisms to hydrolyze peptide

bonds of proteins but, because of evolutionary diversity,

their sizes, specificities and kinetic properties may vary

Medical y interesting proteases in Family C1 (the papain
Papain from the latex of Carica papaya was the first
family) include mammalian enzymes such as cathepsins
sulfhydryl enzyme discovered and has been the subject
B and L (involvement in cancer growth and metastasis)
of mechanism and structural studies for many years
and cathepsin K (of importance for bone degradation an
(Drenth et al., 1971; Glazer and Smith, 1971). The Mr of
osteoporosis) as wel as parasitic enzymes being
papain is 23.4 kDa and pH optimum is 5.5 - 7.0. The
essential for the parasite-host interaction (e.g. cruzipain
enzyme is very stable at neutral pH, even at elevated
from Trypanosoma cruzi - causing Chagas' disease, and
temperatures (Glazer and Smith, 1971). It contains six
falcipain from Plasmodium falciparum - causing malaria).
sulfhydryls and one free cysteine, which is part of the
Predominant expression of cathepsin K in osteoporosis
active site. The complete aminoacid sequence of the
and its wel documented role in bone remodeling makes
enzyme is known, and the three-dimensional structure
cathepsin K and interesting target for the pharmaceutical
has been determined by X-ray crystal ography (Drenth et
industry. Enzymes belonging to Family C13 (the legum-
al., 1971). Schechter and Berger (1967) concluded that
ain family) have been shown to play key roles in antigen
as many as seven sites for recognizing substrate amino
presentation. Interleukin converting enzyme (ICE) and
acid residues exist on the enzyme, al contributing to
other enzymes belonging to Family C14 (the caspase
substrate specificity. It hydrolyses amides of arginine,
family) have gained much interest recently, as key media-
lysine readily and glutamine, histidine, glycine and
tors of apoptosis. Significant activation of calpain, often
tyrosine at reduced rates (Glazer and Smith, 1971).
associated with loss of calcium homeostasis, implicated
Besides papain, papaya latex also contains chymopa-
in pathology of several diseases like muscular dystrophy,
pain, (Jansen and Bal s, 1941) and papaya peptidase A
stroke, traumatic brain injury, alzheimer’s disease, cancer
now known as caricain (Schak et al., 1967). Al the three
and type 2 diabetes mel itus (Carragher, 2006). As a
endopeptidases differ in primary structure, have very
result of recent reports that animal papain-like proteases
similar substrate specificities and are general y assayed
are involved in several pathological conditions, interest in
with synthetic substrates having Arg in P1 position.
the development of inhibitors has substantial y increased.
Another fraction has been detected that demonstrated
Many pharmaceutical companies are seeing it as big
activity against the glycine ester but not against Bz-Arg-
drug market and several clinical trails of inhibitors are
Pna, the ideal substrate for the three endopeptidases
under process (Table 2). To ensure selectivity of inhibi-
isolated from papaya latex. This fraction was named
tion, which is major chal enge, structure based drug
papaya peptidase B. This enzyme is now cal ed
design is becoming more desirable. In addition to animal
proteinase IV or glycyl endopeptidase (Buttle et al.,
sources, structural determination of papain-like proteases
form plant and other sources would certainly provide
The proteolytic enzymes of the pineapple plant Ananas
better understanding of the structural feature of these
comosus are known as bromelains, from stem cal ed
proteases and help us in designing inhibitors for papain-
stem bromelain and from fruit cal ed fruit bromelain.
like proteases for therapeutic application.
There has been a considerable confusion as to whether

these enzymes are distinct proteins (Ota et al., 1972; Ota
et al., 1985) or represent two forms of the same enzyme

(Iida et al., 1973; Sasaki et al., 1973). The pineapple
Plant sources have yielded many useful endopeptidases,
plant has been shown to contain at least 4 distinct endo-
among them calotropins (Abraham and Joshi, 1979), bro-

1082 Afr. J. Biotechnol.

Table 2. Pharmaceutical application of inhibitor of papain-like proteases.

Inhibitor/ clinical trial
Caspase-1 (ICE)
osteoarthritis, Psoriasis
VX-740 / Phase II
Caspase-1 (ICE)
VX-765/ Phase I

Caspase (Broad Spectrum)
Liver diseases
IDN-6556/ Phase IIb

Acute myocardial infarction
2Idun Pharmaceuticals
Caspase-1 (ICE)
Inflammatory (Asthma, arthritis)
IDN-9862, Preclinical
Cathepsin K
SB-462795/ Phase I
Cathepsin K
AAE581/ Phase IIb

1; 2; 3;

peptidases (Rowan et al., 1988; Rowan et al., 1990).
1988; Kaneda et al., 1995). Accordingly, it is presumed
These include, besides stem and fruit bromelain, two
that the bottom of the S2 pocket of melain G is shal ow
other cysteine endopeptidases, ananain (Rowan et al.,
due to the presence of a Phe residue, and a bulky P2
1988) and comosain (Rowan et al., 1990). Similarly,
substrate (for example Phe residue) is not preferred by
fivecysteine proteases known as ficins have been purified
the enzyme. Negatively charged residues at the P3
to homogeneity from the latex of Ficus glabrata (Jones
positions of substrates wel suited the S3 site of melain G
and Glazer, 1970). Al the enzymes had a molecular
for making a salt bridge. Thus, it seems that the sensitive
mass of 25 - 26 kDa with an amino terminal residue
binding pockets of melain G were consequently formed
leucine. They displayed similar specificity and kinetic
by S2 and S3. So far as is known, this is the first reported
properties towards the insulin B chain. Actinidin is an
protease having substrate specificity like this (Uchikoba
anionic protease isolated from the latex of Actinidia
et al., 1999). Also, melain G was little affected by the
chinensis (Chinese gooseberry). The enzyme shows a
inhibitor E-64. It seems that the conformation of E-64 is
molecular mass of 15.4 kDa and is inhibited by DTNB
wel suited for the formation of enzyme-inhibitor complex
and iodoacetamide (McDowal , 1970). The latex of
against the cysteine proteases such as papain, cathepsin
Calotropis gigantea also contain four cysteine proteases
B and calpain, but not for melain G. This also shows that
designated as calotropin FI, FII (Abraham and Joshi,
the subsite of melain G is different from that of papain.
1979a, 1979b) and Calotropin DI, DII (Pal and Sinha,
A cysteine protease of molecular mass 61 kDa has
1980). Two groups of cysteine proteases cal ed asclep-
been identified in the juice of the stem of Dieffenbachia
ains have been isolated from the latex of Asclepias
maculata. The pH optimum is 8.0 and the enzyme is
syriaca and a representative of each has been purified.
inhibited by PCMB and iodoacetate (Chitre et al., 1998).
Asclepains A3 and B5 are homogeneous proteins with
However, protease activity in leaves, petiole and stem
molecular weights of 23 kDa and 21 kDa respectively.
exhibited different pH optima, indicating a possibility that
They are inhibited by thiol specific inhibitors and have a
different molecular forms of protease exist in these parts.
pH optimum of 7.0 - 7.5 (Brockbank and Lynn, 1979).
The enzyme in leaves could be a neutral protease,
Five forms of asclepains have been purified to
whereas alkaline proteases are apparently present in
homogeneity and the sequence of first 21 residues has
petiole and stem. The highest enzyme activity was
been determined and compared to papain (Lynn et al.,
present in the stem. Age of the plant affects the activity
1980a, 1980b).
and the optimum pH of the enzyme. Highest protease
A cysteine protease has been isolated from the ripe
activities were recorded in old (yel owing) leaves, mature
yel ow fruits of the bead tree, Melia azedarach (Kaneda
petiole and mature stem. A cysteine protease with leucyl
et al., 1988). Later, the pressed juice of greenish fruits of
peptidase activity was isolated from stem. A similar
the tree showed very high caseinolytic activity leading to
cysteine protease has been recently isolated from the
the isolation of another cysteine protease cal ed melain G
young stems of Asparagus officinalis, using cystatin
(Uchikoba et al., 1999) and the protease isolated from
affinity chromatography. The molecular mass was
ripe juice was cal ed melain R. From the sites cleaved in
estimated to be 28 kDa and the pH optimum was 7.0
the oxidized insulin B-chain and synthetic oligopeptide
(Yonezawa et al., 1998).

substrates by melain G, the enzyme preferred smal
Similarly, a number of cysteine proteases with novel
amino acid residues such as Gly or Ser at the P2 position
properties have been isolated in our laboratory from the
and negatively charged residues such as glutamic or
latex of Ervatamia coronaria, a flowering plant indigenous
cysteic acid at the P3 position. This is clearly different
to India (Sundd et al., 1998; Kundu et al., 2000; Dubey
from the specificity of papain, which prefers the large
and Jagannadham. 2003a).
hydrophobic amino acid residues such as Phe, Val, and
Being secreted or lysosomal enzymes, peptidases of
Leu at the P2 position (Drenth et al., 1971; Asboth et al.,
the papain family are synthesized with signal peptides,

Dubey et al. 1083

and there are also propeptides at the N-terminus.
ain super family (Bol er, 1986). Cysteine proteases of
Proteolytic cleavage of the propeptides is necessary for
plants play a major role in intracel ular and extra cel ular
activation of the proenzymes. The majority of the prope-
processes such as development and ripening of