Characterization of the guanidine hydrochloride-denatured state of iso-1-cytochrome c by infrared spectroscopy.

Abstract:

:Infrared spectroscopy has been used to monitor residual ordered structure in the denatured state of wild-type and two mutants of iso-1-cytochrome c. The technique used involves a careful digital subtraction procedure that removes spectral contributions from buffer, water vapor, and the denaturant guanidine hydrochloride. Reliable and reproducible spectra can be produced using these methods. The data for iso-1-cytochrome c show upon denaturation a shift of the structure-sensitive amide I infrared band away from the spectral region associated with random structure. Second-derivative resolution enhancement of the amide I absorption band uncovers several bands which can be associated with various residual ordered structures in the denatured state. Gradual changes in the amide I band after denaturation are also observed as the guanidine hydrochloride concentration is increased. Two single-site mutants of iso-1-cytochrome c, which have been shown to have more compact denatured states than the wild-type protein, exhibited denatured-state infrared spectra with significant differences from the wild-type protein spectra. These observations provide new insight into the characteristics of protein denatured states.

journal_name

Biochemistry

journal_title

Biochemistry

authors

Bowler BE,Dong A,Caughey WS

doi

10.1021/bi00175a008

subject

Has Abstract

pub_date

1994-03-08 00:00:00

pages

2402-8

issue

9

eissn

0006-2960

issn

1520-4995

journal_volume

33

pub_type

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