Cofactor-Induced Refolding:  Refolding of Molten Globule Carbonic Anhydrase Induced by Zn(II) and Co(II)

The stability versus unfolding to the molten globule intermediate of bovine carbonic anhydrase II (BCA II) in guanidine hydrochloride (GuHCl) was found to depend on the metal ion cofactor [Zn(II) or Co(II)], and the apoenzyme was observed to be least stable. Therefore, it was possible to find a dena...

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Bibliographic Details
Published in:Biochemistry (Easton) 2001-03, Vol.40 (9), p.2653-2661
Main Authors: Andersson, Dick, Hammarström, Per, Carlsson, Uno
Format: Article
Language:English
Subjects:
Anilino Naphthalenesulfonates - chemistry
Animals
Apoenzymes - chemistry
Binding Sites
Carbonic Anhydrases - chemistry
Cattle
Circular Dichroism
Cobalt - chemistry
Fluorescent Dyes - chemistry
Guanidine - chemistry
Holoenzymes - chemistry
Kinetics
NATURAL SCIENCES
NATURVETENSKAP
Protein Denaturation
Protein Folding
Spectrometry, Fluorescence
Tryptophan
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Summary:The stability versus unfolding to the molten globule intermediate of bovine carbonic anhydrase II (BCA II) in guanidine hydrochloride (GuHCl) was found to depend on the metal ion cofactor [Zn(II) or Co(II)], and the apoenzyme was observed to be least stable. Therefore, it was possible to find a denaturant concentration (1.2 M GuHCl) at which refolding from the molten globule to the native state could be initiated merely by adding the metal ion to the apo molten globule. Thus, refolding could be performed without changing the concentration of the denaturant. The molten globule intermediate of BCA II could still bind the metal cofactor. Cofactor-effected refolding from the molten globule to the native state can be summarized as follows:  (1) initially, the metal ion binds to the molten globule; (2) compaction of the metal-binding site region is then induced by the metal ion binding; (3) a functioning active center is formed; and (4) finally, the native tertiary structure is generated in the outer parts of the protein.
ISSN:0006-2960
1520-4995
1520-4995
DOI:10.1021/bi000957e