Specificity of S '(1) and S '(2) subsites of human tissue kallikrein using the reactive-centre loop of kallistatin: the importance of P '(1) and P '(2) positions in design of inhibitors

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dc.contributor.author Pimenta, Daniel C. [UNIFESP]
dc.contributor.author Fogaca, Sandro E.
dc.contributor.author Melo, Robson L.
dc.contributor.author Juliano, Luiz [UNIFESP]
dc.contributor.author Juliano, Maria Aparecida [UNIFESP]
dc.date.accessioned 2016-01-24T12:33:48Z
dc.date.available 2016-01-24T12:33:48Z
dc.date.issued 2003-05-01
dc.identifier http://dx.doi.org/10.1042/BJ20021952
dc.identifier.citation Biochemical Journal. London: Portland Press, v. 371, p. 1021-1025, 2003.
dc.identifier.issn 0264-6021
dc.identifier.uri http://repositorio.unifesp.br/handle/11600/27217
dc.description.abstract We have demonstrated that the S-1' and S-2', subsites of human tissue kallikrein (hK1) play determinant roles in the recognition and hydrolysis of substrates. the presence of serine at position P-1' and arginine at P-2' resulted in the best substrate, Abz-Ala-Ile-Lys-Phe-Phe-Ser-Arg-Gln-EDDnp, which was derived from the kallistatin reactive-centre loop sequence and quencher groups o-aminobenzoic acid (Abz) and N-(2,4-dinitrophenyl)ethylenediamine (EDDnp). Serine and arginine are also the residues at positions P-1' and P-2' in human kininogen, from which hK1 releases Lys-bradykinin. Several peptide analogues of Abz-Ala-Ile-Lys-Phe-Phe-Ser-Arg-Gln-EDDnp, in which the Ser and Arg residues were substituted with various other amino acids, were synthesized and tested as substrates. Most of them were hydrolysed slowly, although they showed significant binding to hK1, as demonstrated by their competitive inhibition constants (K-i). Using this information, six peptides were designed, synthesized and assayed as inhibitors of hK1. Abz-Lys-Phe-Phe-Pro-Arg-Gln-EDDnp, Abz-Lys-Phe-Arg-Pro-Arg-Gln-EDDnp and acetyl-Lys-Phe-Phe-Pro-Leu-Glu-NH2 inhibited hK1 in the range 20-30 nM (letters in italics denote the D-form of the amino acid). the peptide acetyl-Lys-Phe-Phe-Pro-Leu-Glu-NH2 was a weak inhibitor for other serine proteases, as indicated by the higher K-i values compared with hK1, but this peptide was a potent inhibitor of human plasma kallikrein, which has a Ki value of 8 nM. This result was surprising, since this enzyme is known to be a restricted arginyl-hydrolase. in conclusion, acetyl-Lys-Phe-Phe-Pro-Leu-Glu-NH2 can be used as a leader compound to design specific inhibitors for hK1, plasma kallikrein, or for both at same time, if the inhibition of kinin release is the main goal. en
dc.format.extent 1021-1025
dc.language.iso eng
dc.publisher Portland Press
dc.relation.ispartof Biochemical Journal
dc.rights Acesso aberto
dc.subject bradykinin en
dc.subject inflammation en
dc.subject kallidin en
dc.subject kininogen en
dc.subject serine protease en
dc.title Specificity of S '(1) and S '(2) subsites of human tissue kallikrein using the reactive-centre loop of kallistatin: the importance of P '(1) and P '(2) positions in design of inhibitors en
dc.type Artigo
dc.contributor.institution Universidade Federal de São Paulo (UNIFESP)
dc.description.affiliation Universidade Federal de São Paulo, Escola Paulista Med, Dept Biophys, BR-04044020 São Paulo, Brazil
dc.description.affiliationUnifesp Universidade Federal de São Paulo, Escola Paulista Med, Dept Biophys, BR-04044020 São Paulo, Brazil
dc.identifier.doi 10.1042/BJ20021952
dc.description.source Web of Science
dc.identifier.wos WOS:000182733400040



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