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Substrates and Inhibitors for Proteolytic Enzymes

Grant number: 98/15906-3
Support type:Research Projects - Thematic Grants
Duration: September 01, 1999 - August 31, 2003
Field of knowledge:Biological Sciences - Biochemistry - Enzymology
Principal Investigator:Luiz Juliano Neto
Grantee:Luiz Juliano Neto
Home Institution: Escola Paulista de Medicina (EPM). Universidade Federal de São Paulo (UNIFESP). Campus São Paulo. São Paulo , SP, Brazil

Abstract

The purpose of this project is the development of organic synthesis of peptides and amino acids aimed at the study of proteases. Due to the large diversity of proteolytic enzymes that will be studied and due their relationship with different physiological and physiopathological processes, we will have the collaboration of 6 national and 16 international groups. In addition, will participate in this project 8 graduated students, 5 master students and 3 undergraduate students. Based on our previous experience, we are going to prepare internally quenched fluorescent peptides having as donnor-receptor fluorescent pair ortho-aminobenzoieacid (Abz) and N-[2,4-dinitrophenyll-ethylenediamine (EDDnp), respectively. The following proteolytic enzymes will be studied: tissue and plasma kallikreins, rat tonin, thrombin, cathepsin G, proteases from snake venom, thiol-proteases(cathepsin B, proteases from tropical disease parasites), endooligopeptidases, metalloproteases (angiotensin converting enzyme, 24.11 and 24.15), convertases (Kex2, PCI, PC2,PC5/6 and furin), cathepsin D. Non-natural amino acids will be incorporated in peptides, and modifications on their peptide bond will be performed in order to obtain specific substrates and inhibitors for the proteases we are studying. This project is aimed also at the cell proteolysis, using the substrates and inhibitors that we are developing. In addition, we are proposing to establish a cell culture laboratory for these studies as well as to produce most of the proteases that we are proposing to study. As a consequence of the propositions of this project we will include physical-chemical measurements of some of the obtained peptides in order to correlate form and function of substrates and inhibitors interacting with proteases. Furthermore, other programs are in analysis and planning, which will amplify the present project, namely: a) a systematic study of endo- and exopeptidases from insect digestive tube, looking through an evolutionary perspective, b) proteolysis in germinative processes in plant seeds and/or apoptoses, c) study of kallikreins from human prostate using peptides from reactive loop of natural serpins that inhibited these enzymes, d) capsid protein processing of Dengue virus by viral protease and by convertases of host cells. (AU)