Inactivation, dissociation, and unfolding of tetrameric alcohol dehydrogenase I from Kluyverpmyces lactis (KIADH I) were investigated using guanidinium chloride (GdmCl) as denaturant. Protein transitions were monitored by enzyme activity, intrinsic fluorescence and gel filtration chromatography. At low denaturant concentrations (less than 0.3 M), reversible transformation of enzyme into tetrameric inactive form occurs. At denaturant concentrations between 0.3 and 0.5 M, the enzyme progressively dissociates into structured monomers through an irreversible reaction. At higher denaturant concentrations, the monomers unfold completely. Refolding studies indicate that a total reactivation occurs only with the enzyme denatured between 0 and 0.3 M GdmCl concentrations. The enzyme denatured at GdmC1 concentrations higher than 0.3 M refolds only partially. All together, our results indicate that unfolding of the KIADH I is a multistep process, i.e., inactivation of the structured tetramer, dissociation into partially structured monomers, followed by complete unfolding. (C) 2001 Elsevier Science B.V. All rights reserved.

Multiple unfolded states of alcohol dehydrogenase I from Kluyveromyces lactis by guanidinium chloride / Paolo, Sacchetta; Rosalba Di, Rado; Saliola, Michele; Argante, Bozzi; Falcone, Claudio; Carmine Di, Ilio; Filippo, Martini. - In: BIOCHIMICA ET BIOPHYSICA ACTA-PROTEIN STRUCTURE AND MOLECULAR ENZYMOLOGY. - ISSN 0167-4838. - 1545:1-2(2001), pp. 238-244. [10.1016/s0167-4838(00)00283-1]

Multiple unfolded states of alcohol dehydrogenase I from Kluyveromyces lactis by guanidinium chloride

SALIOLA, Michele;FALCONE, Claudio;
2001

Abstract

Inactivation, dissociation, and unfolding of tetrameric alcohol dehydrogenase I from Kluyverpmyces lactis (KIADH I) were investigated using guanidinium chloride (GdmCl) as denaturant. Protein transitions were monitored by enzyme activity, intrinsic fluorescence and gel filtration chromatography. At low denaturant concentrations (less than 0.3 M), reversible transformation of enzyme into tetrameric inactive form occurs. At denaturant concentrations between 0.3 and 0.5 M, the enzyme progressively dissociates into structured monomers through an irreversible reaction. At higher denaturant concentrations, the monomers unfold completely. Refolding studies indicate that a total reactivation occurs only with the enzyme denatured between 0 and 0.3 M GdmCl concentrations. The enzyme denatured at GdmC1 concentrations higher than 0.3 M refolds only partially. All together, our results indicate that unfolding of the KIADH I is a multistep process, i.e., inactivation of the structured tetramer, dissociation into partially structured monomers, followed by complete unfolding. (C) 2001 Elsevier Science B.V. All rights reserved.
2001
denaturation; dissociation; inactivation; kluyveromyces lactis alcohol dehydrogenase; yeast
01 Pubblicazione su rivista::01a Articolo in rivista
Multiple unfolded states of alcohol dehydrogenase I from Kluyveromyces lactis by guanidinium chloride / Paolo, Sacchetta; Rosalba Di, Rado; Saliola, Michele; Argante, Bozzi; Falcone, Claudio; Carmine Di, Ilio; Filippo, Martini. - In: BIOCHIMICA ET BIOPHYSICA ACTA-PROTEIN STRUCTURE AND MOLECULAR ENZYMOLOGY. - ISSN 0167-4838. - 1545:1-2(2001), pp. 238-244. [10.1016/s0167-4838(00)00283-1]
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11573/96402
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