Determination of disulphide bridges in PG‐2, an antimicrobial peptide from porcine leukocytes

Abstract
We determined the cysteine connectivity of protegrin PG‐2, a leukocyte‐derived antimicrobial peptide, by performing sequential enzyme digestions with chymotrypsin and thermolysin, and monitoring each digest by direct liquid chromatography–electrospray mass spectrometric analysis. This approach resolved the disulphide pairing pattern unambiguously with only picomolar amounts of PG‐2. The inferred cysteine connectivity was confirmed by traditional amino acid composition analyses using nanomolar amounts of the protegrin. The results suggest that protegrins will assume a tachyplesin‐like, disulphide‐stabilized antiparallel β‐sheet configuration in solution.