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Abstract Heterotrophic microorganisms are responsible for transforming and respiring a substantial fraction of the organic matter produced by phytoplankton in the surface ocean. Much of this organic matter is composed of polysaccharides, high‐molecular weight (HMW) sugars. To initiate degradation of polysaccharides, microorganisms must produce extracellular enzymes of the right structural specificity to hydrolyze these complex structures. To date, most measurements of enzyme activities are made at in situ temperatures, but at atmospheric pressure. However, previous studies have shown that hydrostatic pressure can impact the functionality of enzymes. Since deep sea communities may be seeded by microbes from shallow waters, we aimed to determine if pressure affects the performance of enzymes from coastal waters. To determine the extent to which enzymatic activities of coastal microbial communities are affected by pressure, we quantified the degradation of seven polysaccharides under pressures ranging from 0.1 MPa (atmospheric) to 40 MPa (equivalent to 4,000 m). Enzyme activities of pelagic communities were inhibited with increased pressure, while enzyme activities of benthic microbial communities were more resistant to increased pressure. Addition of HMW organic matter resulted in communities with enzyme activities that were more resistant to increased pressure. However, the freely‐dissolved enzymes (<0.2 μm) produced by these communities were strongly inhibited by increased hydrostatic pressure, suggesting that the pressure‐resistant enzymes were cell‐surface attached. Because pressure inhibition of enzyme activities varied strongly by polysaccharide, we surmise that the structural complexity of a polysaccharide—and therefore the number of distinct enzymes required for hydrolysis—is likely closely associated with pressure inhibition.
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On Single-Cell Enzyme Assays in Marine Microbial Ecology and Biogeochemistry
Extracellular enzyme activity is a well-established parameter for evaluating microbial biogeochemical roles in marine ecosystems. The presence and activity of extracellular enzymes in seawater provide insights into the quality and quantity of organic matter being processed by the present microorganisms. A key challenge in our understanding of these processes is to decode the extracellular enzyme repertoire and activities of natural communities at the single-cell level. Current measurements are carried out on bulk or size-fractionated samples capturing activities of mixed populations. This approach – even with size-fractionation – cannot be used to trace enzymes back to their producers, nor distinguish the active microbial members, leading to a disconnect between measured activities and the producer cells. By targeting extracellular enzymes and resolving their activities at the single-cell level, we can investigate underlying phenotypic heterogeneity among clonal or closely related organisms, characterize enzyme kinetics under varying environmental conditions, and resolve spatio-temporal distribution of individual enzyme producers within natural communities. In this perspective piece, we discuss state-of-the-art technologies in the fields of microfluidic droplets and functional screening of prokaryotic cells for measuring enzyme activity in marine seawater samples, one cell at a time. We further elaborate on how this single-cell approach can be used to address research questions that cannot be answered with current methods, as pertinent to the enzymatic degradation of organic matter by marine microorganisms.
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- PAR ID:
- 10338553
- Date Published:
- Journal Name:
- Frontiers in Marine Science
- Volume:
- 9
- ISSN:
- 2296-7745
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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- Free Publicly Accessible Full Text
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Accepted Manuscript1.0
- Journal Article:
- https://doi.org/10.3389/fmars.2022.846656
