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Titolo:
Cystatins as calpain inhibitors: Engineered chicken cystatin- and stefin B-kininogen domain 2 hybrids support a cystatin-like mode of interaction with the catalytic subunit of mu-calpain
Autore:
Diaz, BG; Gross, S; Assfalg-Machleidt, I; Pfeiler, D; Gollmitzer, N; Gabrijelcic-Geiger, D; Stubbs, MT; Fritz, H; Auerswald, EA; Machledit, W;
Indirizzi:
Univ Munich, Adolf Butenandt Inst Physiol Chem Phys Biochem &, D-80336 Munich, Germany Univ Munich Munich Germany D-80336 ys Biochem &, D-80336 Munich, Germany Univ Munich, Klinikum Innenstadt, Chirurg Klin, Abt Klin Chem & Klin Biochem, D-80336 Munich, Germany Univ Munich Munich Germany D-80336 Klin Biochem, D-80336 Munich, Germany Univ La Habana, Fac Biol, Dept Bioquim, La Habana, Cuba Univ La Habana LaHabana Cuba , Fac Biol, Dept Bioquim, La Habana, Cuba Univ Marburg, Inst Pharmazeut Chem, D-35037 Marburg, Germany Univ MarburgMarburg Germany D-35037 zeut Chem, D-35037 Marburg, Germany
Titolo Testata:
BIOLOGICAL CHEMISTRY
fascicolo: 1, volume: 382, anno: 2001,
pagine: 97 - 107
SICI:
1431-6730(200101)382:1<97:CACIEC>2.0.ZU;2-4
Fonte:
ISI
Lingua:
ENG
Soggetto:
CALCIUM-DEPENDENT PROTEASE; RAY CRYSTAL-STRUCTURE; HIGH-LEVEL EXPRESSION; 4 REPEATING DOMAINS; ESCHERICHIA-COLI; CYSTEINE PROTEINASES; CATHEPSIN-B; ENDOGENOUS INHIBITOR; HUMAN CALPASTATIN; PIG CALPASTATIN;
Keywords:
calpastatin; kininogen; papain; stefin B; surface plasmon resonance; temporary inhibition;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Life Sciences
Citazioni:
41
Recensione:
Indirizzi per estratti:
Indirizzo: Machledit, W Univ Munich, Adolf Butenandt Inst Physiol Chem Phys Biochem &, Schillerstr42, D-80336 Munich, Germany Univ Munich Schillerstr 42 MunichGermany D-80336 h, Germany
Citazione:
B.G. Diaz et al., "Cystatins as calpain inhibitors: Engineered chicken cystatin- and stefin B-kininogen domain 2 hybrids support a cystatin-like mode of interaction with the catalytic subunit of mu-calpain", BIOL CHEM, 382(1), 2001, pp. 97-107

Abstract

Within the cystatin superfamily, only kininogen domain 2 (KD2) is able to inhibit mu- and m-calpain. In an attempt to elucidate the structural requirements of cystatins for calpain inhibition, we constructed recombinant hybrids of human stefin B (an intracellular family 1 cystatin) with KD2 and Delta L110 deletion mutants of chicken cystatin-KD2 hybrids. Substitution of the N-terminal contact region of stefin B by the corresponding KD2 sequence resulted in a calpain inhibitor of K-i = 188 nM. Deletion of L110, which forms a beta -bulge in family 1 and 2 cystatins but is lacking in KD2, improved inhibition of mu -calpain 4- to 8-fold. All engineered cystatins were temporary inhibitors of calpain due to slow substrate-like cleavage of a single peptide bond corresponding to Gly9-Ala10 in chicken cystatin. Biomolecular interaction analysis revealed that, unlike calpastatin, the cystatin-type inhibitors do not bind to the calmodulin-like domain of the small subunitof calpain, and their interaction with the mu -calpain heterodimer is completely prevented by a synthetic peptide comprising subdomain B of calpastatin domain 1. Based on these results we propose that (i) cystatin-type calpain inhibitors interact with the active site of the catalytic domain of calpain in a similar cystatin-like mode as with papain and (ii) the potential for calpain inhibition is due to specific subsites within the papain-bindingregions of the general cystatin fold.

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Documento generato il 07/07/20 alle ore 12:28:22