| 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. | |
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MedLine Citation:
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PMID: 15595828 Owner: NLM Status: MEDLINE |
Abstract/OtherAbstract:
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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(1) 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 microM(-1) s(-1)) versus the N-domain (k(cat)/K(m) = 0.51 microM(-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. |
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Authors:
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Patrícia A Bersanetti; Maria Claudina C Andrade; Dulce E Casarini; Maria A Juliano; Aloysius T Nchinda; Edward D Sturrock; Luiz Juliano; Adriana K Carmona |
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Publication Detail:
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Type: Journal Article; Research Support, Non-U.S. Gov't |
Journal Detail:
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Title: Biochemistry Volume: 43 ISSN: 0006-2960 ISO Abbreviation: Biochemistry Publication Date: 2004 Dec |
Date Detail:
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Created Date: 2004-12-14 Completed Date: 2005-02-07 Revised Date: 2006-11-15 |
Medline Journal Info:
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Nlm Unique ID: 0370623 Medline TA: Biochemistry Country: United States |
Other Details:
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Languages: eng Pagination: 15729-36 Citation Subset: IM |
Affiliation:
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Department of Biophysics, Division of Nephrology, Escola Paulista de Medicina, Universidade Federal de São Paulo, Rua 3 de Maio 100, São Paulo 04044-020, Brazil. |
Export Citation:
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| MeSH Terms | |
Descriptor/Qualifier:
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Amino Acid Sequence Angiotensin-Converting Enzyme Inhibitors / chemistry, pharmacology Catalytic Domain Combinatorial Chemistry Techniques / methods* Fluorescence Resonance Energy Transfer / methods Humans Molecular Sequence Data Oligopeptides / chemistry, pharmacology Peptide Library* Peptidyl-Dipeptidase A / chemistry*, drug effects, metabolism* Protein Structure, Tertiary Substrate Specificity |
| Chemical | |
Reg. No./Substance:
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0/Angiotensin-Converting Enzyme Inhibitors; 0/Oligopeptides; 0/Peptide Library; EC 3.4.15.1/Peptidyl-Dipeptidase A |
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