Synthesis and hydrolysis by cathepsin B of fluorogenic substrates with the general structure benzoyl-X-ARG-MCA containing non-natural basic amino acids at position X

Synthesis and hydrolysis by cathepsin B of fluorogenic substrates with the general structure benzoyl-X-ARG-MCA containing non-natural basic amino acids at position X

Author Melo, R. L. Google Scholar
Pozzo, RCB Google Scholar
Alves, L. C. Google Scholar
Perissutti, E. Google Scholar
Caliendo, G. Google Scholar
Santagada, V Google Scholar
Juliano, L. Google Scholar
Juliano, M. A. Google Scholar
Institution Universidade Federal de São Paulo (UNIFESP)
Univ Naples Federico 2
Abstract We synthesized one series of fluorogenic substrates for cathepsin B derived from the peptide Bz-F-R-MCA (Bz = benzoyl, MCA = 7-methyl-coumarin amide) substituting Phe at the P(2) position by non-natural basic amino acids that combine a positively charged group with aromatic or aliphatic radicals at the same side chain, namely, 4-aminomethyl-phenylalanine, 4-guanidine-phenylalanine. 4-aminomethyl-N-isopropyl-phenylalanine. 3-pyridyl-alanine, 4-piperidinyl-alanine, 4-amino-methyl-cyclohexyl-alanine. 4-aminocyclohexyl-alanine, and N(im)-dimethyl-histidine. Bz-F-R-MCA was the best substrate for cathepsin B but also hydrolyzed Bz-R-R-MCA with lower efficiency, since the protease accepts Arg at St due to the presence of Glu(245) at the bottom of this subsite. the presence of the basic non-natural amino acids at the Pt position of the substrate partially restored the catalytic efficiency of cathepsin B. All the kinetic parameters for hydrolysis of the peptides described in this paper are in accordance with the structures of the St pocket previously described. in addition, the substrate with 4-aminocyclohexyl-alanine presented the highest affinity to cathepsin B although the peptide was obtained from a mixture of cis/trans isomers of the amino acid and we were not able to separate them. for comparison all the obtained substrates were assayed with cathepsin L and papain. (C) 2001 Elsevier Science B.V. All rights reserved.
Keywords cathepsin B
cathepsin L
papain
3-aminomethyl-phenylalanine
4-guanidine phenylalanine
3-aminomethyl-N-isopropylphenylalanine
3-pyridyl-alanine
4-piperidinyl-alanine
4-aminomethyl-cyclohexyl-alanine
4-aminocyclohexyl-alanine
N(im)-dimethylhistidine
protease
Language English
Date 2001-05-05
Published in Biochimica Et Biophysica Acta-protein Structure and Molecular Enzymology. Amsterdam: Elsevier B.V., v. 1547, n. 1, p. 82-94, 2001.
ISSN 0167-4838 (Sherpa/Romeo, impact factor)
Publisher Elsevier B.V.
Extent 82-94
Origin http://dx.doi.org/10.1016/S0167-4838(01)00171-6
Access rights Closed access
Type Article
Web of Science ID WOS:000168698100009
URI http://repositorio.unifesp.br/handle/11600/26549

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