Rapid mass spectrometric determination of disulfide connectivity in peptides and proteins

• DOI: 10.1039/c3mb25534d
• Publication Date: 2013-02-08T09:37:35Z
• Authors: Moitrayee Bhattac...; Kallol Gupta; Konkallu Hanumae ...; Padmanabhan Balaram
• Keywords: Oxidative folding; Fragmentation; Folding (DSP implementation); Proteolytic enzymes

Disulfide crosslinks are ubiquitous in natural peptides and proteins, providing rigidity to polypeptide scaffolds. The assignment of disulfide connectivity multiple crosslinked systems is often difficult achieve. Here, we show that rapid unambiguous characterisation can be achieved through direct mass spectrometric CID fragmentation the intact polypeptides. method requires a native bonded polypeptides subsequent analysis using newly developed program, DisConnect. Technical difficulties involving proteins surmounted by an initial proteolytic nick determination structures these While fragments containing one cystine evident from MS profile alone, those with cystines subjected fragmentation. wide applicability this illustrated examples peptide hormones, toxins, foldamers synthetic analogue marine toxin. method, coupled DisConnect, provides unambiguous, straightforward approach, especially useful for screening crosslink fidelity recombinant linkages toxins characterization folding intermediates encountered oxidative pathways.