Abstract
Proteases are enzymes which catalyzes the irreversible hydrolysis of peptide bonds in proteins. Cysteine cathepsins belonging to proteases have also been termed as papain-like protease because they resemble the overall fold of papain. The present chapter aims to focus on the historical aspects, structure, cellular distribution, biosynthesis, mechanism of catalysis, its regulation, physiological functions and its association with rheumatoid arthritis. As these enzymes are also new therapeutic drug targets, information on available assays of cysteine cathepsins and their inhibitors are also highlighted which will help in development of therapies in various diseases.
Key Words : Cysteine cathepsin ''' Rheumatoid arthritis ''' Cystatin ''' Lysosomal proteases ''' Activity regulation ''' Activity based probes ''' Substrates based probes
1. Introduction
The health of an organism is maintained by number of factors, of which the major contribution is governed by controlled biosynthesis, maturation, function and terminal catabolism of proteins. The selective and well controlled proteolytic events are accomplished by a group of proteases. These simple destructive enzymes likely arose at the earliest stages of protein evolution in primitive organisms [1]. The proteases constitute 1-4 % of the genes per genomes sequenced to date [2] and are found in all known life forms from viruses to mammalian. In mammal, two main groups of cysteine proteases [CP] are present: cytosolic proteases (calpains type I and calpains type II) and lysosomal cathepsins [3]. More than 670 proteases and 200 endogenous protease were encoded by the human genome [2,4]. The cysteine proteases also known as cysteine cathepsin [CC] accounts for about 25 % of the proteases and traditionally were believed to perform non specific bulk proteolysis within lysosomes. Cathepsin, derived from the Greek word '''KATHEPSIN''' meaning '''to digest''', was first proposed by Willstatter and Bamann [5] for the acid proteinase activity found in the aqueous extracts of several mammalian tissues.
The lysosomes being discovered more than 50 years, are membrane-bound organelles representing the important degradative compartments in eukaryotic cells. Some cell types, from the hematopoietic lineage, contain a specialized lysosomal compartment which secretes its contents into the extracellular environment in response to external stimuli [6]. These are termed as '''secretory lysosomes''' or lysosomal'''related organelles sharing properties which are similar and differing from the conventional lysosomes. The acidic pH and presence of degradative proteins are found in both types of lysosomes but the secretory lysosomes have the ability to undergo regulated secretion [7].
In this chapter, the main aim is to review the various aspects of CC pertaining to its historical heritage, cellular localization, structure, biosynthesis, mechanism of catalysis, biological functions, structure'''function relationship, activity regulation, available assays, therapeutic strategies and its association with diseased state such as rheumatoid arthritis [RA] to give a current knowledge on CC.
2. Historical heritage
The discovery of lysosomal cysteine proteases also known as cathepsins was in the first half of the 20th century [8]. Since its discovery, it has been known that these enzymes non-selectively degrades proteins in lysosomes and are also responsible for a number of important cellular processes. The term cathepsin comprises proteases from three different mechanistic classes ''' the cysteine, serine and aspartic proteases. In humans, eleven CC are known : cathepsin B,C (J, Dipeptidyl peptidase I or DPPI), F,H,K (O2) , L,O,S,V (L2), X (P,Y,Z) and W (lymphopain) [9]. Being members of papain family, they belong to clan CA (cathepsin) of cysteine proteases. The papain-like thiol protease form the largest subfamily (C1) among the cysteine protease clan CA.
Cathepsin C the first pure enzyme, was found in the 1940s [10]. The enzymes cathepsin B and H were identified in the early 1970s followed soon after by cathepsin L [11].
In early 1980s, the amino acid sequences of rat cathepsins B and H were published. [12].The crystal structure of human cathepsin B was determined in 1990 [13] indicating a rapid progress in this field. It is also known as golden era of lysomal cysteine protease research wherein 6 out of 11 known human enzymes identified.
3. Cellular localization
The physiological function of cathepsins can be partly attributed to the differences in localization of them in specific cell types. The expression profile of cathepsins B, H, L, C, X, F, O and V in human tissues are ubiquitous indicating that these enzymes are involved in a normal cellular protein degradation and turnover [14]. On the other hand, cathepsins K,W and S shows a restricted cell or tissue specific distribution revealing their more specific role. Cathepsin K is highly expressed in osteoclasts, epithelial cells and in synovial fibroblast in RA joints [15] therefore reflecting the major role for cathepsin K in bone resorption. Cathepsin W is predominantly expressed in CD8+ lymphocytes and natural killer cells [16] but its biological function still remains elusive. The antigen-presenting cells (APCs) such as dendritic cells, macrophages and B-cells express cathepsin S [17]. There is high homology between cathepsin L and V but the former cellular expression is ubiquitous in contrast to restricted distribution of the latter to thymus and testis [18]. There is huge body of evidence that active cathepsins are also localized in other cellular compartments such as nucleus, cytoplasm and plasma membrane. Nuclear cathepsin L plays a role in the regulation of cell cycle progression through proteolytic processing of the CDP/Cux transcription factor [19]. The interaction between nucleosomes and stefin B inhibits nuclear cathepsin L there by delaying cell cycle progression [20]. Cathepsin L mediates the proteolysis of histone H3 which modulates cell-cycle progression [21].
4. Structure
The crystal structure of papain [22] together with actinidin [23] has provided the first insights into the three dimensional (3D) structure of cysteine cathepsins. They are small monomeric proteins with molecular weight around 30 kDa with the exception of cathepsin C which is tetrameric with 200 kDa. The CC structurally similar to papain [24] consists of two domains the L- and R- which specify the left and right domain of the catalytic unit and is in accordance with standard view as shown in (Fig. 1).] The L-domain has three '' '''helices, one of which is vertical and also known as central helix having around 30 residues. The R-domain is a kind of ''-barrel with the front strand forming a coiled structure. At the bottom the barrel is enclosed by ''-helix. The two domains separate at the top forming a V-shaped active cleft. In the middle of this cleft the catalytic residues cys 25 from the L-domain and His 159 from the R-domain resides. The proteolytic activity of the enzymes is mediated through these catalytic residues forming thiolate-imidazolium ion pair and has a low pka around 2.5'''3.5 [25].
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