Control of L-malate metabolism in Escherichia coli ML308 (ATCC 15224)

Escherichia coli ML308 (ATCC 15224) grows aerobically on L-malate as a sole source of carbon and energy in a medium otherwise containing only inorganic ions. Cells grow immediately on L-malate if previously grown on other dicarboxylic acids, some and no acids or ketoglutaric acid but lag before growth on L-Malate after growth on glucose, glycerol, pyruvate or acetate. Limitation of growth rate must be due to a factor which is present in cells during growth on the first but not on the second group of carbon sources. The three factors considered were PEP carboxy kinase, malic enzymes and the L-malate transport system. Acetate stimulates growth rate on L-malate in all circumstances and must, therefore, increase the rate of production of all intermediates. In particular the rate of supply of phosphoenol-pyruvate (PEP) for glucogenesis etc. Must be increased. Because PEP is made from L-malate by PEP carboxy kinase it follows that this enzyme cannot limit rate of growth on L-malate alone. The relationship between activities of malic enzymes and growth rate on L-malate was measured directly. Different phenotypes, able to metabolise L-malate at various rates, were found to contain more malic enzyme activity than was expressed during growth on L-malate. Furthermore, malic enzyme activities during adaptation to L-malate do not alter. Consequently, growth rate on L-malate cannot be limited by malic enzyme activities. Maleate has been reported as a gratuitous inducer of the L-Malate transport system. Its presence causes adaptation to L-malate during growth on glycerol, suggesting that transport activity is related to the rate of metabolism of L-malate. The activity of the L-Malate transport system, as measured by the rato of pyruvate production, in arsenite inhibited whole cells, increases with adaptation so confirming that the rate of metabolism of L-malate is limited by its transport. The regulation of central metabolism and the characteristics of the enzymes metabolising L-malate are also consistent with the transport mechanism being the process which limits metabolism. Degree of adaptation was measured as a range of growth rates with corresponding rates of oxygen uptake on L-malate. The relationship between the two permits a maintenance coefficient and YG value to be calculated for L-malate growth. Analogous values are also given for growth on acetate. The full interpretation of these data depends on assumptions as to the efficiency (P/O ratio) of oxidative phosphorylation. It seems probable that P/O ratios and maintenance vary with the growth substrate and that when a lower efficiency of oxidative phosphorylation is obtained, a higher level of maintenance is required. Molar growth yields (as Yc-source and YO2) reflect both P/O ratios and the whole mechanism of biosynthesis. Consideration of the apparent YATP for different carbon sources strongly suggests that the P/O ration varies and depends on the carbon source available to the cells. It is not surprising that the highest P/O ratios are obtained with those carbon sources which support the highest growth rates