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Titolo:
Functional properties of the active core of human cystathionine beta-synthase crystals
Autore:
Bruno, S; Schiaretti, F; Burkhard, P; Kraus, JP; Janosik, M; Mozzarelli, A;
Indirizzi:
Univ Parma, Inst Biochem Sci, I-43100 Parma, Italy Univ Parma Parma Italy I-43100 a, Inst Biochem Sci, I-43100 Parma, Italy Univ Parma, Natl Inst Phys Matter, I-43100 Parma, Italy Univ Parma ParmaItaly I-43100 tl Inst Phys Matter, I-43100 Parma, Italy Univ Basel, Biozentrum, ME Muller Inst Struct Biol, CH-4056 Basel, Switzerland Univ Basel Basel Switzerland CH-4056 ct Biol, CH-4056 Basel, Switzerland Univ Colorado, Sch Med, Dept Pediat, Denver, CO 80262 USA Univ Colorado Denver CO USA 80262 Med, Dept Pediat, Denver, CO 80262 USA
Titolo Testata:
JOURNAL OF BIOLOGICAL CHEMISTRY
fascicolo: 1, volume: 276, anno: 2001,
pagine: 16 - 19
SICI:
0021-9258(20010105)276:1<16:FPOTAC>2.0.ZU;2-O
Fonte:
ISI
Lingua:
ENG
Soggetto:
O-ACETYLSERINE SULFHYDRYLASE; TRYPTOPHAN SYNTHASE; ALPHA(2)BETA(2) COMPLEX; SALMONELLA-TYPHIMURIUM; HEME PROTEIN; 3-DIMENSIONAL STRUCTURE; PYRIDOXAL 5'-PHOSPHATE; ENZYME; MICROSPECTROPHOTOMETRY; BINDING;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Life Sciences
Citazioni:
35
Recensione:
Indirizzi per estratti:
Indirizzo: Mozzarelli, A Univ Parma, Inst Biochem Sci, I-43100 Parma, Italy Univ Parma Parma Italy I-43100 m Sci, I-43100 Parma, Italy
Citazione:
S. Bruno et al., "Functional properties of the active core of human cystathionine beta-synthase crystals", J BIOL CHEM, 276(1), 2001, pp. 16-19

Abstract

Human cystathionine beta -synthase is a pyridoxal 5'-phosphate enzyme containing a heme binding domain and an S-adenosyl-L-methionine regulatory site. We have investigated by single crystal microspectrophotometry the functional properties of a mutant lacking the S-adenosyhmethionine binding domain. Polarized absorption spectra indicate that oxidized and reduced hemes are reversibly formed. Exposure of the reduced form of enzyme crystals to carbon monoxide led to the complete release of the heme moiety. This process, which takes place reversibly and without apparent crystal damage, facilitatesthe preparation of a heme-free human enzyme. The heme-free enzyme crystalsexhibited polarized absorption spectra typical of a pyridoxal 5'-phosphate-dependent protein. The exposure of these crystals to increasing concentrations of the natural substrate L-serine readily led to the formation of the key catalytic intermediate alpha -aminoacrylate. The dissociation constant of L-serine was found to be 6 mM, close to that determined in solution. Theamount of the alpha -aminoacrylate Schiff base formed in the presence of L-serine was pH independent between 6 and 9, However, the rate of the disappearance of the alpha -aminoacrylate, likely forming pyruvate and ammonia, was found to increase at pH values higher than 8, Finally, in the presence of homocysteine the alpha -aminoacrylate-enzyme absorption band readily disappears with the concomitant formation of the absorption band of the internal aldimine, indicating that cystathionine beta -synthase crystals catalyze both beta -elimination and beta -replacement reactions. Taken together, these findings demonstrate that the heme moiety is not directly involved in the condensation reaction catalyzed by cystathionine beta -synthase.

ASDD Area Sistemi Dipartimentali e Documentali, Università di Bologna, Catalogo delle riviste ed altri periodici
Documento generato il 28/09/20 alle ore 23:39:42