Molecular modeling of the substrate specificity of prohormone convertases SPC2 and SPC3.
In this paper we describe the results of molecular modeling of the structures of the active sites of two subtilisin-like prohormone convertases (SPCs), SPC2 (PC2) and SPC3 (PC1/PC3). These enzymes are members of a recently discovered family of cellular proteases involved in the processing of precursor proteins. Although these proteases all possess catalytic domains similar to the bacterial subtilisins no tertiary structural data from x-ray analysis are yet available. We have shown that despite the high structural homology of the subtilisins and the SPCs, the structure of the loop which lies immediately below the active sites differs due to the presence of a cis-peptide bond (Tyr167-Pro168) in this loop in the subtilisins and its absence in the SPCs. Accordingly, we have proposed a new alignment for the amino acid sequences of the SPCs in this region. Both SPC2 and SPC3 participate in the processing of prohormones at dibasic cleavage sites, typically Lys-Arg or Arg-Arg. To investigate the structural basis of the substrate specificity of these SPCs, we have carried out molecular mechanic calculations of the optimal arrangement and interactions of peptide substrates containing several residues of arginine or lysine, i.e. Arg, Ala-Ala-Ala-Arg, Arg-Ala-Ala-Arg, Arg-Ala-Arg-Arg, Arg-Ala-Lys-Arg, in the putative active sites. Such subtilisin-based modeling has allowed us to identify those negatively charged residues, Asp and Glu, in the S1, S2, and S4 subsites, which can directly interact with basic residues in the substrates via formation of salt bridges and thereby contribute to the substrate selectivity of the SPCs.
Lipkind
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Department of Biochemistry and Molecular Biology, University of Chicago, Illinois 60637, USA.
Gong
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Steiner
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DF
D000595:Amino Acid Sequence; D016282:Aspartic Acid Endopeptidases / Q000737:chemistry / Q000378:metabolism*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D045706:Proprotein Convertase 2; D043484:Proprotein Convertases; D011487:Protein Conformation; D013379:Substrate Specificity; D013381:Subtilisins / Q000737:chemistry / Q000378:metabolism*
D043484:Proprotein Convertases; D013381:Subtilisins; D045706:Proprotein Convertase 2; D016282:Aspartic Acid Endopeptidases