Open AccessDetermination of disulfide bridges in natural and recombinant insect defensin A
Author(s) -
LEPAGE Pierre,
BITSCH Francis,
ROECKLIN Dominique,
KEPPI Elisabeth,
DIMARCQ JeanLuc,
REICHHART JeanMarc,
HOFFMANN Jules A.,
ROITSCH Carolyn,
DORSSELAER Alain
Publication year - 1991
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1991.tb15872.x
Subject(s) - edman degradation , chemistry , recombinant dna , mass spectrometry , defensin , chromatography , peptide , protein primary structure , disulfide bond , thermolysin , biochemistry , peptide sequence , enzyme , trypsin , gene
The primary‐structure comparison of natural insect defensin A from Phormia terranovae and recombinant insect defensin A from Saccharomyces cerevisiae has been accomplished using a combination of Edman degradation and liquid secondary ion mass spectrometry. The natural and recombinant proteins have the same primary structure with identical disulfide‐bond designations ( and ) as determined from the peptides obtained after thermolysin digestion. The combined use of Edman degradation and mass spectrometry allowed the disulfide‐bridge structure to be determined with a total of only 40μg (9.9 nmol) natural peptide. Mass spectrometry provides a rapid means of disulfide‐bridge verification, requiring not more than 20μg recombinant insect defensin A, which is compatible with use in batch analysis.