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(Reference retrieved automatically from Web of Science through information on FAPESP grant and its corresponding number as mentioned in the publication by the authors.)

Substrate specificity of mitochondrial intermediate peptidase analysed by a support-bound peptide library

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Author(s):
Marcondes, M. F. M. [1] ; Alves, F. M. [1] ; Assis, D. M. [1] ; Hirata, I. Y. [1] ; Juliano, L. [1] ; Oliveira, V. [1] ; Juliano, M. A. [1]
Total Authors: 7
Affiliation:
[1] Univ Fed Sao Paulo, Escola Paulista Med, Dept Biophys, Sao Paulo - Brazil
Total Affiliations: 1
Document type: Journal article
Source: FEBS OPEN BIO; v. 5, p. 429-436, 2015.
Web of Science Citations: 1
Abstract

The substrate specificity of recombinant human mitochondrial intermediate peptidase (hMIP) using a synthetic support-bound FRET peptide library is presented. The collected fluorescent beads, which contained the hydrolysed peptides generated by hMIP, were sequenced by Edman degradation. The results showed that this peptidase presents a remarkable preference for polar uncharged residues at P-1 and P-1' substrate positions: Ser = Gln > Thr at P-1 and Ser > Thr at P-1'. Non-polar residues were frequent at the substrate P-3, P-2, P-2' and P-3' positions. Analysis of the predicted MIP processing sites in imported mitochondrial matrix proteins shows these cleavages indeed occur between polar uncharged residues. Previous analysis of these processing sites indicated the importance of positions far from the MIP cleavage site, namely the presence of a hydrophobic residue (Phe or Leu) at P-8 and a polar uncharged residue (Ser or Thr) at P-5. To evaluate this, additional kinetic analyses were carried out, using fluorogenic substrates synthesized based on the processing sites attributed to MIP. The results described here underscore the importance of the P-1 and P-1' substrate positions for the hydrolytic activity of hMIP. The information presented in this work will help in the design of new substrate-based inhibitors for this peptidase. (C) 2015 The Authors. Published by Elsevier B.V. on behalf of the Federation of European Biochemical Societies. This is an open access article under the CC BY-NC-ND license. (AU)

FAPESP's process: 12/50191-4 - Synthesis, kinetic studies and applications of substrates and inhibitors for proteolytic enzymes
Grantee:Maria Aparecida Juliano
Support Opportunities: Research Projects - Thematic Grants
FAPESP's process: 11/20941-9 - DEVELOPMENT AND USE OF FLUORESCENCE TECHNIQUES FOR THE STUDY OF PROTEIN CONFORMATIONAL CHANGES
Grantee:Marcelo Ferreira Marcondes Machado
Support Opportunities: Scholarships in Brazil - Post-Doctoral
FAPESP's process: 14/00661-0 - Development and use of fluorescence techniques for the study of protein conformational changes
Grantee:Marcelo Ferreira Marcondes Machado
Support Opportunities: Scholarships abroad - Research Internship - Post-doctor