Ubiquitin function studied by disulfide engineering.

D J Ecker; T R Butt; J Marsh; E Sternberg; A Shatzman; J S Dixon; P L Weber; S T Crooke

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PubMed · 2536376

Ubiquitin function studied by disulfide engineering.

D J Ecker·T R Butt·J Marsh·E Sternberg·A Shatzman·J S Dixon·P L Weber·S T Crooke

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Abstract

Disulfide engineering was used to probe the role of conformational mobility in ubiquitin-mediated proteolysis. Six genes that encode cysteine-containing mutants of ubiquitin were constructed, expressed in Escherichia coli and the proteins purified. Single cysteine-containing mutants and a 4/14 disulfide were active in degradation of a substrate protein in vitro, while the 4/66 disulfide, which cross-links the NH2- and COOH-terminal strands of the protein, was only 20-30% active. The solution structure of the 4/66 mutant was solved: the disulfide is left-handed with no perturbations in the backbone from that of wild type ubiquitin. The results suggest that conformational mobility is required for the activity of ubiquitin in signaling proteolysis.

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Computer Simulation Cystine Disulfides Humans Magnetic Resonance Spectroscopy Mutation Protein Conformation Ubiquitins Journal Article

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D J Ecker, T R Butt, J Marsh, E Sternberg, A Shatzman, J S Dixon, P L Weber, S T Crooke. 1989-01-25. Ubiquitin function studied by disulfide engineering.. https://pubmed.ncbi.nlm.nih.gov/2536376/

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