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 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.Universidade Federal de São Paulo, Dept Biophys, Div Nephrol, Escola Paulista Med, BR-04044020 Observatory, SP, BrazilUniversidade Federal de São Paulo, Dept Med, Div Nephrol, Escola Paulista Med, BR-04044020 São Paulo, BrazilUniv Cape Town, Div Med Biochem, Inst Infect Dis & Mol Med, ZA-7925 Cape Town, South AfricaUniversidade Federal de São Paulo, Dept Biophys, Div Nephrol, Escola Paulista Med, BR-04044020 Observatory, SP, BrazilUniversidade Federal de São Paulo, Dept Med, Div Nephrol, Escola Paulista Med, BR-04044020 São Paulo, BrazilWeb of Scienc

Living in the waterfalls: A new species of Trichomycterus (Siluriformes: Trichomycteridae) from Tabay stream, Misiones, Argentina.

A new species assigned to the genus Trichomycterus from the area of the waterfalls of Tabay stream, Paraná River basin, Misiones, Argentina, is described. Trichomycterus ytororo sp. nov. is distinguished from all other species in the genus by the presence of 31-35 dorsal procurrent caudal-fin rays and the combination of some external characters such as: coloration, number of pectoral-fin rays and pores of the laterosensory canals. The new taxon belongs to a presumably monophyletic group of species composed of T. crassicaudatus, T. igobi, and T. stawiarski based on the presence of 24 or more thickly ossified and rigid procurrent caudal-fin rays with a slender distal tip extending along the tips of at least ten neural spines