Rapid Analysis of Single-Cysteine Variants of Recombinant Proteins
Protein engineering methods have been widely used to study individual structural factors that contribute to protein stability (1,2). An important goal of that research is to enhance the commercial or medicinal utility of wild-type (WT) proteins by increasing then stability in a rational, step-by-step fashion (3). We recently characterized a series of single-cysteine variants of subtilisin BPN′, a proteolytic enzyme used in commercial laundry formulations. They had been prepared (4) in part because random mutagenesis experiments indicated that some of these variants were more stable than the WT enzyme (3). We wanted to compare the stabilities of the mutants with those observed after engineering single disulfide bonds into the subtilisin BPN′ backbone (5,6).
KeywordsTryptic Peptide Sinapinic Acid MALDI Mass Spectrum Surface Accessibility Tryptic Fragment
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