A Computational Tool to Detect and Avoid Redundancy in Selected Reaction Monitoring□S

Selected reaction monitoring (SRM), also called multiple reaction monitoring, has become an invaluable tool for tar-geted quantitative proteomic analyses, but its application can be compromised by nonoptimal selection of transi-tions. In particular, complex backgrounds may cause am-biguities in SRM measurement results because peptides with interfering transitions similar to those of the target peptide may be present in the sample. Here, we developed a computer program, the SRMCollider, that calculates nonredundant theoretical SRM assays, also known as unique ion signatures (UIS), for a given proteomic back-ground. We show theoretically that UIS of three transitions suffice to conclusively identify 90%of all yeast peptides and 85%of all human peptides. Using predicted retention times, the SRMCollider also simulates time-scheduled SRMacqui-sition, which reduces the number of interferences to con-sider and leads to fewer transitions necessary to construct an assay. By integrating experimental fragment ion intensi-ties from large scale proteome synthesis efforts (SRMAtlas) with the information content-based UIS, we combine two orthogonal approaches to create high quality SRM assays ready to be deployed. We provide a user friendly, open source implementation of an algorithm to calculate UIS of any order that can be accessed online a