Bioinformatics of granzymes: sequence comparison and structural studies on granzyme family by homology modeling.
Cytotoxic lymphocytes (CTLs), the key players of cell mediated immunity, induce apoptosis by engaging death receptors or through exocytosis of cytolytic granules containing granzyme (proteases) and pore-forming protein (perforin). The crystal structure of granzyme B from human (B(h)) and rat (B(r)), as well as that of pro-granzyme K (K(h)) has been reported recently. In the present communication, we describe the homology modeling of granzyme family (in particular Gzm A(h), M(h), B(m), and C(m) from human and mouse) based on the crystal structural coordinates of trypsin, granzyme K (K(h)), and granzyme B (B(h)). These models have been used for establishing phylogenetic relationship as well as identifying characteristic features for designing specific inhibitors. The paper also highlights key residues at the S1, S2, and S2(') binding subsites in all granzyme, which may be involved in the structure-function relationship of this enzyme family. The predicted 3D homology models show a conserved two similar domain structure, i.e., an N-terminal domain and a C-terminal domain comprising predominantly of beta-sheet structure with a little alpha-helical content. Micro-heterogeneities have been observed in the vicinity of the active site in all granzymes as compared to granzyme B(h). For example, in granzyme M(h), valine is present at the S1 subsite instead of arginine. Similarly differences at S2 (Leu-->Phe), S3 (Ser-->Gly), and S4 (Arg-->Asn) subsites are quite apparent and appear to hold the potential for selective designing of inhibitors for possible therapeutic applications. Furthermore, analysis of the electrostatic surface potential on the shape of granzyme-inhibitor binding groove reveals clear differences at the reactive site. Additionally the different posttranslational modification sites such as phosphorylation (e.g., in granzyme M Thr101, Ser109), myristoylation (Gly22, 117, and 131), and glycosylation (Ser160) have been identified, as very little is known about the functional significance of these modifications in the granzyme family. Thus, glycosylation at Ser160 in granzyme M may influence the net charge of the enzyme, resulting in altered substrate binding as compared to granzyme B. Also this modification may influence the rate of complexation and binding affinity with proteoglycans. These studies are expected to contribute towards the basic understanding of functional associations of the granzymes with other molecules and their possible role in apoptosis.
Sattar
Rabia
R
International Center for Chemical Sciences, HEJ Research Institute of Chemistry, University of Karachi, Karachi 75270, Pakistan.
Ali
S Abid
SA
Abbasi
Atiya
A
D000595:Amino Acid Sequence; D000818:Animals; D017209:Apoptosis; D001665:Binding Sites; D019295:Computational Biology; D053804:Granzymes; D006801:Humans; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010802:Phylogeny; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D051381:Rats; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D012697:Serine Endopeptidases / Q000737:chemistry*; D013329:Structure-Activity Relationship; D013379:Substrate Specificity; D013602:T-Lymphocytes, Cytotoxic / Q000378:metabolism; D014357:Trypsin / Q000737:chemistry; D053802:Tryptases
C509190:Tpsb2 protein, mouse; C509213:GZMB protein, human; C509218:GZMM protein, human; D053804:Granzymes; C509214:Gzmb protein, mouse; C509215:Gzmb protein, rat; C509216:Gzmc protein, mouse; C509217:Gzmc protein, rat; C509219:Gzmm protein, mouse; C509220:Gzmm protein, rat; D012697:Serine Endopeptidases; D014357:Trypsin; C509222:Tpsab1 protein, mouse; C509223:Tpsab1 protein, rat; C509221:Tpsb2 protein, rat; D053802:Tryptases; C509212:GZMA protein, human