A free flow electrophoresis separation strategy for segregation of high abundant phycobilisomes from cyanobacterium nostoc punctiforme PCC 73102

Abstract

Generally proteomics analysis of cyanobacteria has limited dynamic resolution because of the presence of abundant proteins. Thus proteomics assessment of the low abundance proteins in cyanobacteria is difficult as it comprises phycobilisome and Rubisco. The current investigation assesses the performance of FFE in isoelectric focusing (IEF) mode: a solution-based technique that separates proteins on the basis of isoelectric point (pI). We explore the advantages of combining robust protein fractionation using FFE and 1D-LC-MS/MS to extend protein identification deeper through the dynamic range of the model cyanobacterium, Nostoc punctiforme PCC 73102. Sixty-one novel new proteins (out of 248 (all identified with ≥ 2 peptides)) were successfully identified using FFE-IEF compared to all previous reports. Results demonstrate the ability of FFE to provide improved protein distributions, while also providing effective segregation of highly abundant phycobilisomes to allow access to 37 low abundance proteins (pI > 9) in the N. punctiforme proteome, which are difficult to observe using conventional methods. © 2012 Wase NV, et al.