CysMap and CysJoin: Database and tools for protein disulphides localisation

Abstract

We have developed a computer program able to make user-customised databases derived from the public PIR nonredundant reference protein database. When the database of interest has been created, the user will generate the map of all the possible linear peptides containing one and two cysteines for each protein and combine them to calculate the mass of all the possible clusters of linear peptides linked by a disulphide bridge with a cysteine pair. It is also possible to create selected maps corresponding to peptides formed by the action of specific proteases. In this way, mass spectrometric data obtained from the hydrolysis of proteins of unknown sequence can be related to that contained in the database for quick disulphide assignment and protein identification. To confirm signal attribution, the program will also furnish the expected mass of cluster peptides after performing a cycle of Edman degradation. The utility of the program is discussed and examples of application are given.

High‐throughput method for on‐target performic acid oxidation of MALDI‐deposited samples

“…Disulfide bonds play important roles in stabilizing and maintaining native protein structure,1, 2 and mass spectrometry (MS) plays an increasingly important role in deciphering these processes,3 especially protein folding/unfolding kinetics 4–6. Disulfide bond locations within proteins can be determined using a variety of MS‐based methods, including tandem MS,7, 8 database search algorithms,9–12 chemical reduction,13–15 chemical oxidation16–18 and metal ion cleavage of disulfide bonds 19, 20. Xu and coworkers developed a tandem MS search engine to simplify database searches for disulfide‐linked peptides fragment ion spectra,12 but such spectra can be complicated by direct cleavage of the disulfide bond between interlinked disulfide peptides, which results in a limited number of sequence‐informative fragment ions from each α‐ and β‐chain peptide forming the disulfide linkage 7.…”