Kinetic analysis of spermine binding to NRD convertase

Kinetic analysis of spermine binding to NRD convertase

Autor Csuhai, E. Google Scholar
Juliano, M. A. Google Scholar
Juliano, L. Google Scholar
Hersh, L. B. Google Scholar
Instituição Univ Kentucky
Universidade Federal de São Paulo (UNIFESP)
Resumo N-arginine dibasic convertase cleaves polypeptides between paired basic residues containing the sequence Arg-Arg or Arg-Lys. the enzyme contains a large anionic domain, which in the rat enzyme consists of 57 acidic residues out of a stretch of 76 amino acids. Polyamines modulate the activity of the enzyme presumably by binding at the anionic domain (Csuhai ct al. (1995) Biochemistry 34, 12411-12419). in this study a kinetic analysis of the effect of salts and amines, particularly the polyamine spermine, on the rat enzyme was studied, Simple salts were inhibitory with no apparent specificity for the anion or cation. Inhibition resulted in an increased K-m and a decreased V-max. Evidence that amines bind to an anionic domain was obtained by the finding that N,N-bis [2-hydroxyethyl]-2-aminoethanesulfonic acid, which is structurally related to the inhibitory amine triethanolamine, is noninhibitory. Inhibition exhibited a complex dependence on spermine concentration. the data fit a model in which enzyme-spermine and enzyme-(spermine), complexes are formed. A pH-independent K-d (similar to 0.1 mu M) was obtained for enzyme-spermine formation, while enzyme-(spermine), formation was dependent on pH; K-d at pH 6.5 = 1 mu M and a K-d at pH 8 = similar to 16 mu M. Direct binding of spermine was demonstrated by the ability of spermine to increase the thermal stability of the enzyme. the concentration dependence for the spermine-induced increase in thermal stability fits a model in which formation of the enzyme-spermine complex is sufficient to account for the observed changes. (C) 1999 Academic Press.
Idioma Inglês
Data 1999-02-15
Publicado em Archives of Biochemistry and Biophysics. San Diego: Academic Press Inc, v. 362, n. 2, p. 291-300, 1999.
ISSN 0003-9861 (Sherpa/Romeo, fator de impacto)
Editor Academic Press Inc
Extensão 291-300
Direito de acesso Acesso restrito
Tipo Artigo
Web of Science WOS:000078675200013

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