Expression of glutamine synthetase and glutamate dehydrogenase by marine bacterioplankton: Assay optimizations and efficacy for assessing nitrogen to carbon metabolic balance in situ

Matthew P. Hoch, Richard A. Snyder, Wade H. Jeffrey, Kevin S. Dillon,Richard B. Coffin

Limnol. Oceanogr. Methods 4:308-328 (2006) | DOI: 10.4319/lom.2006.4.308

ABSTRACT: Expression of glutamate metabolism enzymes, glutamate dehydrogenase (GDH), and glutamine synthetase (GS) are proposed to yield information on the nitrogen (N) to carbon (C) metabolic balance within bacterioplankton communities. Whole-cell assay conditions were optimized, and reactions were linear with time and biomass. Enzyme activities were assayed in seawater cultures from four ecosystems of contrasting trophic state amended with different regimes of glucose, amino acids, and ammonium (NH4+) to validate expression patterns of GDH and GS in various combinations of N and C limitation or excess. In three of four experiments, glucose amendment enhanced GS expression by 2-fold but repressed GDH by 10% to 40% relative to the control. In contrast, addition of amino acids or NH4+ resulted in 20% to 90% repression of GS and enhanced GDH expression by 20% to 900%. The GDH:GS activity ratio (×10–3) ranged from 6 to 22 in glucose added treatments and 63 to 264 in NH4+-amended treatments and appears to be a more sensitive index of bacterial N bioavailability relative to C supply than either enzyme alone. Cluster analysis was used to identify the condition of ambient bacterioplankton by matching enzyme expression with the validation results. This enzymatic approach overcomes some of the biases and limitations of other methods for assessing N metabolism in marine bacteria while providing unique information to further understand constraints of bacterioplankton respiration, production, and N flux.