Context dependence of mutational effects in a protein: The crystal structures of the V35I, I47V and V35I/I47V gene V protein core mutants

Hong Zhang, Matthew M. Skinner, Warren S. Sandberg, Andrew H J Wang, Thomas C. Terwilliger

Research output: Contribution to journalArticle

20 Scopus citations

Abstract

The basis for the context dependence of the effects of core mutations on protein stability was investigated by comparing the structures of three gene V protein mutants with that of the wild-type protein. We previously examined a 'swapped' mutant in which core residues Val35 and Ile47 were simply reversed so that the mutant had no hydrophobicity change from the native protein. The swapped mutant was destabilized by 3 kcal/mol per gene V protein dimer relative to the wild-type protein, demonstrating that factors other than hydrophobicity must make substantial contributions to the effects of mutations on the stability of the protein. Here we have determined the structure of this swapped mutant (V35I/I47V) as well as those of the two constituent mutants (V35I and I47V). We find that the structures of the mutant proteins are very similar to that of the wild-type protein except for the necessary addition or deletion of methylene groups and for slight positional shifts of atoms around each mutated residue. The structure of the double mutant is a composite of the structures of the two single mutants. In the mutant structures, the V35I mutation fills a cavity that exists in the wild-type protein and the I47V mutation creates a new cavity. The structures of the mutants indicate further that the reason the V35I and I47V mutations do not have opposite effects on stability is that the cavity in the wild-type protein filled by the V35I mutation is not optimally shaped for accommodating the additional methylene group of the isoleucine. These results support the concepts that the details of core packing have substantial influence on the effects of core mutations on protein stability and that these packing effects are major determinants of the context dependence of core mutation effects on stability.

Original languageEnglish (US)
Pages (from-to)148-159
Number of pages12
JournalJournal of Molecular Biology
Volume259
Issue number1
DOIs
StatePublished - May 31 1996

Keywords

  • Filamentous phage
  • Gene V protein
  • Protein stability
  • Site-specific mutants
  • X-ray crystallography

ASJC Scopus subject areas

  • Structural Biology
  • Molecular Biology

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