Guanidine hydrochloride denaturation studies of mutant forms of staphylococcal nuclease.

Research output: Contribution to journalArticle

Abstract

Several mutant forms of staphylococcal nuclease with one or two defined amino acid substitutions have been purified, and the effects of the altered amino acid sequence on the stability of the folded conformation have been analyzed by guanidine hydrochloride denaturation. Two nuc- mutations, which greatly reduced the level of enzyme activity accumulated in E coli colonies carrying a recombinant plasmid with the mutant nuc gene (ie, a NUC- phenotype), both result in protein unfolding at significantly lower guanidine hydrochloride concentrations than the wild-type protein, whereas three sup mutations isolated on the basis of their ability to suppress partially the NUC- phenotype of the above two mutations result in unfolding at significantly higher guanidine hydrochloride concentrations. Characterization of nuclease molecules with two different amino acid substitutions, either nuc- + sup pairs or sup + sup pairs, suggests that the effect of an amino acid substitution on the stability of the native conformation, as measured by the value of delta delta GD, may not be a constant, but rather a variable that is sensitive to the presence of other substitutions at distant sites in the same molecule. Surprisingly, the slopes of the log Kapp vs guanidine hydrochloride concentration plots vary by as much as 35% among the different proteins.

Original languageEnglish (US)
Pages (from-to)281-289
Number of pages9
JournalJournal of Cellular Biochemistry
Volume30
Issue number4
StatePublished - 1986

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Micrococcal Nuclease
Denaturation
Guanidine
Substitution reactions
Amino Acid Substitution
Amino Acids
Mutation
Conformations
Phenotype
Protein Unfolding
Proteins
Molecules
Enzyme activity
Escherichia coli
Amino Acid Sequence
Plasmids
Genes
Enzymes

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology

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Guanidine hydrochloride denaturation studies of mutant forms of staphylococcal nuclease. / Shortle, David R.

In: Journal of Cellular Biochemistry, Vol. 30, No. 4, 1986, p. 281-289.

Research output: Contribution to journalArticle

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