Accommodation of single amino acid insertions by the native state of staphylococcal nuclease

John Sondek, David Shortle

Research output: Contribution to journalArticlepeer-review


Single alanine and glycine insertions were introduced at 20 randomly selected positions in staphylococcal nuclease. The resulting changes in catalytic activity and in stability to guanidine hydrochloride denaturation indicate that the native state structure is frequently able to accommodate the extra residue without great difficulty, even insertions within secondary structural elements such as alpha helices and beta sheets. On average, an inserted residue reduces the free energy of denaturation (ΔG H 2O) by an amount roughly comparable to an alanine or glycine substitution for one of the residues flanking the site of insertion. Several positions outside of the enzyme active site were found where insertions, but not substitutions, lead to structural changes that modify catalytic activity and the circular dichroism spectrum. Amino acid insertions represent a virtually unexplored class of genetic mutation that may prove complementary to amino acid substitutions for engineering proteins with altered functional and structural properties.

Original languageEnglish (US)
Pages (from-to)299-305
Number of pages7
JournalProteins: Structure, Function, and Bioinformatics
Issue number4
StatePublished - 1990


  • alanine insertion
  • circular dichroism spectra
  • conformational changes
  • glycine insertion
  • guanidine hydrochloride denaturation
  • polypeptide backbone
  • protein stability

ASJC Scopus subject areas

  • Structural Biology
  • Biochemistry
  • Molecular Biology


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