Stabilization of myoglobin by multiple alanine substitutions in helical positions

Laura Lin, Rachel J. Pinker, George N. Phillips, Neville R. Kallenbach

Research output: Contribution to journalArticlepeer-review


We have carried out a series of multiple Xaa → Ala changes at nonadjacent surface positions in the sequence of sperm whale myoglobin. Although the corresponding single substitutions do not increase the thermal stability of the protein, multiple substitutions enhance the stability of the resulting myoglobins. The effect observed is an increase in the observed Tm (midpoint unfolding temperature) relative to that predicted from assuming additivity of the free energy changes corresponding to single mutations. The stabilization occurs in the presence of urea, as measured by the dependence of the unfolding temperature on urea concentration. The sites that have been altered occur in different helices and are not close in sequence or in the native structure of myoglobin. The observed effect is consistent with a role of multiple alanines in residual interactions in the unfolded state of the mutant proteins.

Original languageEnglish (US)
Pages (from-to)1430-1435
Number of pages6
JournalProtein Science
Issue number9
StatePublished - Sep 1994


  • protein folding
  • protein stability
  • sperm whale myoglobin
  • α‐helix

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology


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