Throughout the world’s oceans, about half of the carbon dioxide that is fixed by photosynthetic microbes annually (~50 gigatons) is transferred into seawater as dissolved organic matter (DOM). This pool of fixed carbon is typically made up of a mixture of sugars, proteins, nucleic acids and other organic molecules. Marine microbes are particularly well adapted to using this complex energy source, since they produce specialized enzymes to digest and transporter proteins to take up these dissolved compounds. Considering the immense size of this resource pool and the important role of microbes in its digestion, the turnover of DOM by microbes in marine environments can significantly affect many large-scale processes, such as global climate patterns and the transfer of energy through marine food webs. Although we have gained much knowledge about the genetic diversity and potential activity of marine bacteria, the links between their identity and the substrates they are breaking down are not well understood. Our research is focused on developing techniques to address this question, by attempting to define which specific microbial populations use different types of fixed carbon, and how different microbes may work together to break down large molecules, or if they carry out these processes in relative isolation from one another.
Specifically, our project aims to link the identity of marine heterotrophic bacteria with the organic substrates that they incorporate into biomass with two relatively new stable isotope probing (SIP) methods. We will use Chip-SIP (NanoSIMS analysis of phylogenetic microarrays) to determine the 16S phylotypes incorporating major types of DOM molecules (lipid, protein, sugars, etc.) and originating from major phytoplankton taxa (diatoms, cyanobacteria, dinoflagellates). Concurrently, we will use proteomics-SIP to examine the metabolic incorporation patterns of the carbon substrates into newly synthesized proteins. Microbial community diversity and functional potential of samples collected from the coastal Pacific Ocean will be first defined using metagenomic approaches. This information will be used in combination with Chip-SIP and proteomics-SIP methods to differentiate the active and inactive community members during SIP labeling experiments incubated over a short period of time in the presence of the 13C-labeled molecules defined above. Our results will provide a direct link between coastal heterotrophic bacterial populations and the origin (phytoplankton species) and molecular form (size and major type) of the organic carbon that sustains their growth.
Ryan S. Mueller and Chongle Pan (2013) Sample handling and mass spectrometry for microbial metaproteomic analyses. Methods in Enzymology, Volume 531, Pages 289-303
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