Deciphering disulfide bond patterns in proteins remains a significant challenge. In the present study, interlinked disulfide bonds connecting peptide chains are homolytically cleaved with 193 nm ultraviolet photodissociation (UVPD). Analysis of insulin showcased the ability of UVPD to cleave multiple disulfide bonds and provide sequence coverage of the peptide chains in the same MS/MS event. For proteins containing more complex disulfide bonding patterns, an approach combining partial reduction and alkylation mitigated disulfide scrambling and allowed assignment of the array of disulfide bonds. The 4 disulfide bonds of lysozyme and the 19 disulfide bonds of serotransferrin were characterized through LC/UVPD-MS analysis of nonreduced and partially reduced protein digests.
Deciphering disulfide bond patterns in proteins remains a significant challenge. In the present study, interlinked n class="Chemical">disulfide bonds connecting peptide chains are homolytically cleaved with 193 nm ultraviolet photodissociation (UVPD). Analysis of insulin showcased the ability of UVPD to cleave multiple disulfide bonds and provide sequence coverage of the peptide chains in the same MS/MS event. For proteins containing more complex disulfide bonding patterns, an approach combining partial reduction and alkylation mitigated disulfide scrambling and allowed assignment of the array of disulfide bonds. The 4 disulfide bonds of lysozyme and the 19 disulfide bonds of serotransferrin were characterized through LC/UVPD-MS analysis of nonreduced and partially reduced protein digests.
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