Examination of Microbial Proteome Preservation Techniques Applicable to Autonomous Environmental Sample Collection

Improvements in temporal and spatial sampling frequency have the potential to open new windows into the understanding of marine microbial dynamics. In recent years, efforts have been made to allow automated samplers to collect microbial biomass for DNA/RNA analyses from moored observatories and autonomous underwater vehicles. Measurements of microbial proteins are also of significant interest given their biogeochemical importance as enzymes that catalyze reactions and transporters that interface with the environment. We examined the influence of five preservatives solutions (SDS-extraction buffer, ethanol, trichloroacetic acid, B-PER, and RNAlater) on the proteome integrity of the marine cyanobacterium Synechococcus WH8102 after four weeks of storage at room temperature. Four approaches were used to assess degradation: total protein recovery, band integrity on an SDS-PAGE gel, and number of protein identifications and relative abundances by 1D LC-MS/MS proteomic analyses. Total protein recoveries from the preserved samples were lower than the frozen control due to processing losses, which could be corrected for with internal standardization. The trichloroacetic acid preserved sample showed significant loss of protein band integrity on the SDS-PAGE gel. The RNAlater preserved sample showed the highest number of protein identifications (103% relative to the control; 520 + 31 identifications in RNAlater versus 504 + 4 in the control), equivalent to the frozen control. Relative abundances of individual proteins in the RNAlater treatment were quite similar to that of the frozen control (average ratio of 1.01 + 0.27 for the 50 most abundant proteins), while the SDS-extraction buffer, ethanol, and B-PER all showed significant decreases in both number of identifications and relative abundances of individual proteins. Based on these findings, RNAlater was an effective proteome preservative, although further study is warranted on additional marine microbes.