Chemical modification of bovine liver rhodanese with tetrathionate: differential effects or the sulfur-free and sulfur-containing catalytic intermediates

A. R.S. Prasad, Paul M. Horowitz

Research output: Contribution to journalArticlepeer-review

7 Scopus citations


Sulfhydryl groups of bovine liver rhodanese (thiosulfate: cyanide sulfurtransferase, EC were modified by treatment with tetrathionate. There was a linear relationship between loss of enzyme activity and the amount of tetrathionate used. At a ratio of one tetrathionate per mole of rhodanese, 100% of enzyme activity was lost in the sulfur-free E-form as compared with a 70% loss for the sulfur-containing ES-form of the enzyme. Addition of up to a 100-fold molar excess of tetrathionate to ES gave no further inactivation. Addition of cyanide to the maximally inactivated ES-tetrathionate complex gave complete loss of activity. Kinetic studies of maximally inactivated ES and partially inactivated E gave Km (K5) values that were essentially the same as native enzyme, indicating that the active enzyme, in all cases, bound thiosulfate-similarly. Reactivation was faster with the ES-form than with the E-form. The substrate, thiosulfate, could reactivate the enzyme up to 70% in 1 h with ES as compared to 24 h with E. Tetrathionate modification of rhodanese could be correlated with the changes in intrinsic fluorescence and with the binding of the active site reporter 2-anilinonaphthalene-8-sulfonic acid (2,8-ANS). Circular dichroism spectra of the protein suggested increased ordered secondary structure in the protein after reaction with tetrathionate. Cadmium chloride and phenylarsine oxide totally inactivated the enzyme at levels usually associated with their effect on enzymes containing vicinal sulfhydryl groups. Further, cadmium inhibition could be reserved by EDTA. Tetrathionate modification of rhodanese may proceed through the formation of sulfenylthiosulfate intermediates at sulfhydryl groups, close to but not identical with the active-site sulfhydryl group, which then can react further with the active-site sulfhydryl group to form disulfide bridges.

Original languageEnglish (US)
Pages (from-to)102-108
Number of pages7
JournalBiochimica et Biophysica Acta (BBA)/Protein Structure and Molecular
Issue number1
StatePublished - Jan 5 1987
Externally publishedYes


  • Chemical modification
  • Enzyme inactivation
  • Rhodanese
  • sulfhydryl group
  • Tetrathionate

ASJC Scopus subject areas

  • Biophysics
  • Structural Biology
  • Biochemistry
  • Molecular Biology


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