Beasley J R, Pielak G J
Department of Chemistry, University of North Carolina at Chapel Hill 27599-3290, USA.
Proteins. 1996 Sep;26(1):95-107. doi: 10.1002/(SICI)1097-0134(199609)26:1<95::AID-PROT9>3.0.CO;2-F.
Cassette mutagenesis was used to produce a library of mutations at the interface of the N- and C-terminal helices of Saccharomyces cerevisiae iso-1-cytochrome c. The library is random and comprises > 98% mutations. Over 11,000 candidates were assayed for function by selecting for the ability of yeast, with the mutated gene as their sole cytochrome c source, to grow on nonfermentable carbon sources. We estimate that approximately 0.5% of the 160,000 total amino acid combinations at these four residues result in a functional cytochrome c. Significant correlations are found between the phenotype of yeast harboring the alleles and both the Dayhoff mutation matrix and transfer free energies (cyclohexane-to-water and n-octanol-to-water). Similar correlations are observed with respect to growth rate. Finally, sequences that are consistent with function follow a binary amino acid pattern.
