Interactive intermediates are formed during the urea unfolding of rhodanese

P. M. Horowitz, M. Butler

Resultado de la investigación: Articlerevisión exhaustiva

36 Citas (Scopus)

Resumen

Structural transitions have been studied on the pathway for urea denaturation of rhodanese. Unlike guanidinium hydrochloride, urea gives no visible precipitation. Increasing urea concentrations cause a transition in which the enzyme activity is completely lost by 4.5 M urea, and there is a shift of the intrinsic fluorescence maximum from 335 nm for the native enzyme to 350 nm. There is a maximum exposure of organized hydrophobic surfaces at 4.5 M urea as reported by the fluorescence of 1,1'-bi(4-anilino)naphthalene- 5,5'-disulfonic acid. Above 4.5 M urea, this probe reports the progressive loss of organized hydrophobic surfaces. The polarization of the intrinsic fluorescence falls with increasing urea concentrations in a complex transition showing that rhodanese flexibility increases in at least two phases. Rhodanese becomes increasingly susceptible to digestion by subtilisin between 3.5 and 4.5 M urea, giving rise to large fragments. At urea concentrations >5 M, rhodanese is completely digested. There is a small increase in the rate of sulfhydryl accessibility between 3.5 and 4.5 M urea, but there is a large increase in the sulfhydryl accessibility above 4.5 M urea. Dimethyl suberimidate cross-linking shows the presence of associated species in 3-5 M urea, but there are few cross-linkable species at lower or higher urea concentrations. These results are consistent with a model in which urea unfolding of rhodanese is associated with the initial production of a species having organized regions of structure with exposed hydrophobic surfaces separated by flexible elements.

Idioma originalEnglish (US)
Páginas (desde-hasta)2500-2504
Número de páginas5
PublicaciónJournal of Biological Chemistry
Volumen268
N.º4
EstadoPublished - 1993
Publicado de forma externa

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Huella

Profundice en los temas de investigación de 'Interactive intermediates are formed during the urea unfolding of rhodanese'. En conjunto forman una huella única.

Citar esto