Determination of the disulfide array in the human defensin HNP-2. A covalently cyclized peptide.
HNP-2 is a 29-residue peptide present in human neutrophils and is a member of the defensin family of antimicrobial peptides. All defensins contain an invariant disulfide infrastructure comprised of 6 half-cystine residues. The disulfide structure of HNP-2 was determined using a novel method to identify the cross-links involving the amino- and carboxyl-terminal cysteine residues. A derivative of HNP-2 was synthesized by covalent modification of the terminal cysteine residues. This derivative was purified, characterized, and subjected to exhaustive proteolytic digestion. Characterization of purified proteolytic fragments by amino acid analysis and/or sequence analysis identified an oligopeptide containing all 6 cystine residues. This oligopeptide was subjected to a single cycle of Edman degradation to cleave the peptide bond linking 2 adjacent cysteines. Purification and characterization of the Edman reaction products allowed for assignment of the disulfide array in HNP-2, revealing a cystine motif unique to the defensin peptide family. Further, the covalent structure of HNP-2 was found to be cyclic as one disulfide links the amino- and carboxyl-terminal cysteine residues. HNP-2 is the only polypeptide known to possess such a configuration.
Selsted
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ME
Department of Medicine, University of California, Los Angeles 90024.
Harwig
S S
SS
D000595:Amino Acid Sequence; D001798:Blood Proteins / Q000648:ultrastructure*; D003553:Cystine; D004220:Disulfides; D006801:Humans; D008969:Molecular Sequence Data; D009504:Neutrophils / Q000502:physiology*; D010446:Peptide Fragments / Q000032:analysis; D010449:Peptide Mapping; D023084:alpha-Defensins
D001798:Blood Proteins; D004220:Disulfides; D010446:Peptide Fragments; D023084:alpha-Defensins; C406416:human neutrophil peptide 2; D003553:Cystine