Shotgun proteomics: Tools for analysis of marine particulate proteins
Limnol. Oceanogr. Methods 7:865-874 (2009) | DOI: 10.4319/lom.2009.7.865
ABSTRACT: This study sought a high resolution and high-throughput method to identify and characterize proteins from marine particulate organic matter (POM) using proteomic approaches. The results showed that only a limited number of discrete protein spots were distinguished using two-dimensional electrophoresis (2-DE). Most protein spots were faint and small in 2-DE gels, with a heavy unresolved smeared staining background, indicating 2-DE was not a good high resolution method to separate particulate proteins for identification and characterization. The shotgun proteomic approach combining one-dimensional electrophoresis and capillary liquid chromatography-tandem mass spectrometry as well as the NCBI protein database search was successfully applied to identify and characterize particulate proteins. Using this approach, 737 proteins matching one or more peptides were detected in a POM sample collected from the 41 m water layer in the basin area of the western South China Sea. Of these, 184 were identified as high-confidence proteins matching two or more peptides, including photosynthetic proteins, transporters, molecular chaperones, and porins. In addition to these proteins with known functions, a significant number of novel proteins (accounting for ~30% of the proteins identified) were also detected. The identification of a large number of high-confidence proteins in the POM sample demonstrated that the shotgun proteomic approach is reliable and feasible for the study of particulate proteins and will provide a powerful tool to comprehensively investigate the nature and dynamics of POM in the ocean.