Positional-scanning combinatorial libraries of fluorescence resonance energy transfer peptides for defining substrate specificity of the angiotensin I-converting enzyme and development of selective C-domain substrates

Positional-scanning combinatorial libraries of fluorescence resonance energy transfer peptides for defining substrate specificity of the angiotensin I-converting enzyme and development of selective C-domain substrates

Autor Bersanetti, P. A. Google Scholar
Andrade, MCC Google Scholar
Casarini, D. E. Google Scholar
Juliano, M. A. Google Scholar
Nchinda, A. T. Google Scholar
Sturrock, E. D. Google Scholar
Juliano, L. Google Scholar
Carmona, A. K. Google Scholar
Instituição Universidade Federal de São Paulo (UNIFESP)
Univ Cape Town
Resumo Positional-scanning combinatorial libraries of fluorescence resonance energy transfer peptides were used for the analyses of the S-3 to S-1' subsites of the somatic angiotensin I-converting enzyme (ACE). Substrate specificity of ACE catalytic domains (C- and N-domains) was assessed in an effort to design selective substrates for the C-domain. Initially, we defined the S, specificity by preparing a library with the general structure Abz-GXXZXK(Dnp)-OH [Abz = o-aminobenzoic acid, K(Dnp) = N-epsilon-2,4-dinitrophenyllysine, and X is a random residue], where Z was successively occupied with one of the 19 natural amino acids with the exception of Cys. the peptides containing Arg and Leu in the P-1 position had higher C-domain selectivity. in the sublibraries Abz-GXXRZK(Dnp)-OH, Abz-GXZRXK(Dnp)-OH, and Abz-GZXRXK(Dnp)-OH, Arg was fixed at P-1 so we could define the C-domain selectivity of the S-1', S-2, and S-3 subsites. On the basis of the results from these libraries, we synthesized peptides Abz-GVIRFK(Dnp)-OH and Abz-GVILFK(Dnp)-OH which contain the most favorable residues for C-domain selectivity. Systematic reduction of the length of these two peptides resulted in Abz-LFK(Dnp)-OH, which demonstrated the highest selectivity for the recombinant ACE C-domain (k(cat)/K-m = 36.7 muM(-1) s(-1)) versus the N-domain (k(cat)/K-m = 0.51 muM(-1) s(-1)). the substrate binding of Abz-LFK(Dnp)-OH with testis ACE using a combination of conformational analysis and molecular docking was examined, and the results shed new light on the binding characteristics of the enzyme.
Idioma Inglês
Data de publicação 2004-12-21
Publicado em Biochemistry. Washington: Amer Chemical Soc, v. 43, n. 50, p. 15729-15736, 2004.
ISSN 0006-2960 (Sherpa/Romeo, fator de impacto)
Publicador Amer Chemical Soc
Extensão 15729-15736
Fonte http://dx.doi.org/10.1021/bi048423r
Direito de acesso Acesso restrito
Tipo Artigo
Web of Science WOS:000225776200008
Endereço permanente http://repositorio.unifesp.br/handle/11600/28052

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