Microbial protein in soil: influence of extraction method and C amendment on extraction and recovery.
The capacity to study the content and resolve the dynamics of the proteome of diverse microbial communities would help to revolutionize the way microbiologists study the function and activity of microorganisms in soil. To better understand the limitations of a proteomic approach to studying soil microbial communities, we characterized extractable soil microbial proteins using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Two methods were utilized to extract proteins from microorganisms residing in a Quitman and Benfield soil: (1) direct extraction of bulk protein from soil and (2) separation of the microorganisms from soil using density gradient centrifugation and subsequent extraction (DGC-EXT) of microbial protein. In addition, glucose and toluene amendments to soil were used to stimulate the growth of a subset of the microbial community. A bacterial culture and bovine serum albumin (BSA) were added to the soil to qualitatively assess their recovery following extraction. Direct extraction and resolution of microbial proteins using SDS-PAGE generally resulted in smeared and unresolved banding patterns on gels. DGC-EXT of microbial protein from soil followed by separation using SDS-PAGE, however, did resolve six to 10 bands in the Benfield but not the Quitman soil. DGC-EXT of microbial protein, but not direct extraction following the addition of glucose and toluene, markedly increased the number of bands (approximately 40) on the gels in both Benfield and Quitman soils. Low recoveries of added culture and BSA proteins using the direct extraction method suggest that proteins either bind to soil organic matter and mineral particles or that partial degradation takes place during extraction. Interestingly, DGC may have been preferentially selected for actively growing cells, as gauged by the 10-100x lower cy19:0/18:1omega7 ratio of the fatty acid methyl esters in the isolated community compared to that for the whole soil. DGC can be used to isolate soil communities and provide microbial protein that can be characterized using PAGE.
Plant and Soil Sciences, Mississippi State University, Starkville, MS, USA.
Erin B
EB
Taylor
Mark A
MA
Williams
D001419:Bacteria / Q000254:growth & development / Q000302:isolation & purification; D001426:Bacterial Proteins / Q000302:isolation & purification*; D002499:Centrifugation, Density Gradient; D004591:Electrophoresis, Polyacrylamide Gel; D005947:Glucose / Q000737:chemistry; D015999:Multivariate Analysis; D020543:Proteome; D040901:Proteomics / Q000379:methods*; D012710:Serum Albumin, Bovine / Q000737:chemistry; D012987:Soil / Q000032:analysis*; D012988:Soil Microbiology; D014050:Toluene / Q000737:chemistry
D001426:Bacterial Proteins; D020543:Proteome; D012987:Soil; D012710:Serum Albumin, Bovine; D014050:Toluene; D005947:Glucose