RESPIRATION OF PSEUDOMONAS-FLUORESCENS AS A FUNCTION OF INTRACELLULARSUBSTRATE CONCENTRATION

A kinetic method to measure the intracellular concentration of respira tory substrates in short-term starvation-enrichment experiments is pro posed. Samples of bacterial suspension from steady-state chemostat cul tures were subjected to 25 min starvation, followed by pulse addition of [C-14]glucose. Residual substrate utilization rates and respiration rates (uptake of dissolved O-2) before and after amendment were recor ded. Increases in pool sizes (Delta L) during transients were calculat ed on the basis of C balance. The dependence of respiration rate q(res p) on Delta L was found to obey modified Michaelis-Menten kinetics: q( resp) = Q(resp)((L) over cap+Delta L)/(K-L+(L) over cap+Delta L) [Q(re sp) is maximal respiration rate (29.1 mmol O-2 h(-1) per g biomass C), K-L = 12.14 mg C per g biomass C], where (L) over cap is the absolute value of the pool size before amendment. Direct chemical determinatio n of (L) over cap in cold TCA extracts revealed two fractions. The fir st fraction was mobile and showed a close correlation with both respir ation and L. The second, 'stable', fraction did not correlate with res piration dynamics and was interpreted as material formed artifactually by acid degradation of polymeric cell components.