Characterization of lysine acetylation in the peripheral subunit-binding domain of the E2 subunit of the pyruvate dehydrogenase-2-oxoglutarate dehydrogenase hybrid complex from Corynebacterium glutamicum

ABSTRACT In Corynebacterium glutamicum, pyruvate dehydrogenase (PDH) and 2-oxoglutarate dehydrogenase (ODH) form a unique hybrid complex in which CgE1p and CgE1o are associated with the CgE2–CgE3 subcomplex. We analyzed the role of a lysine acetylation site in the peripheral subunit-binding domain of CgE2 in PDH and ODH functions. Acetylation-mimic substitution at Lys391 of CgE2 severely reduced the interaction of CgE2 with CgE1p and CgE3, but not with CgE1o, indicating the critical role of this residue in the assembly of CgE1p and CgE3 into the complex. It also suggested that Lys391 acetylation inhibited the binding of CgE1p and CgE3 to CgE2, thereby affecting PDH and ODH activities. Interestingly, the CgE2-K391R variant strain showed increased l-glutamate production and reduced pyruvate accumulation. Kinetic analysis suggested that the increased affinity of the K391R variant toward pyruvate might be advantageous for l-glutamate production.

Na+-dependent HCO3-transport in the cyanobacteriumSynechocystisPCC6803

The effect of Na+on HCO3-transport, inorganic carbon (Ci) accumulation, and photosynthesis was investigated in the unicellular cyanobacterium Synechocystis sp. PCC6803 using the silicone fluid filtering centrifugation technique. Unlike other cyanobacteria, Synechocystis cells grown at low Ciin standing culture had little capacity for Na+-independent HCO3-transport, when assayed at pH 9.6. However, 25 mM NaCl, but not KCl, strongly promoted HCO3-transport and accumulation. Kinetic analysis indicated that the HCO3-concentration required for one half the maximum rate of transport, K0.5(HCO3-), decreased in the presence of Na+while the maximum rate of transport, VMAX, increased by up to 15-fold. Na+-dependent HCO3-transport occurred against an electrochemical potential of up to 24 kJ ·mol-1, indicating the involvement of carrier-mediated active transport. Li+(1-3 mM) partially substituted for Na+in that K0.5( HCO3-) values were similar (38 vs. 50 µM), but VMAXwas reduced by twofold. At higher concentrations, Li+counteracted the effects of Na+. Monensin reversibly inhibited Na+-dependent HCO3-transport and acted by reducing VMAXwithout affecting K0.5(HCO3-). Monensin inhibition suggested that the electrochemical potential for Na+may play a role in Na+-dependent HCO3-transport, possibly through an involvement in intracellular pH regulation during transport. Na+also stimulated photosynthetic C fixation and O2evolution and these effects were correlated with the Na+-dependent increase in intracellular Ciaccumulation. The Na+-requirement for photosynthesis could be relieved by the provision of CA to the cell suspension, in agreement with the proposal that Na+is required for transport and not directly involved in the photosynthetic process.Key words: active transport, CO2-concentrating mechanism, cyanobacteria, Na+-dependent HCO3-transport, photosynthesis, Synechocystis PCC6803.

Metabolism of Ferulic Acid Sucrose Esters in Anthers of Tulipa cv. Apeldoorn: I. The Accumulation of Esters and Free Sugars

The authors of Tulipa cv. Apeldoorn accumulate a large amount of ferulic acid sucrose esters. In addition to the well-known diferuloyl-(DFS) and triferuloyl sucrose esters (TFS), a new phenylpropanoid characterized as a monoester of ferulic acid and sucrose is described. Like T F S and D F S , this ester exhibits an accumulation m a x i m u m in the early stages of pollen development. Numerous free sugars have been extracted from Tulipa anthers. Three of them were identified as sucrose, glucose and fructose. Sucrose as the main component is accumulated in extremely high amounts during specific developmental stages (= 20 % of the dry weight). After separation of the anthers into an anther wall fraction (AWF) and pollentapetum (PTF), each fraction shows a different accumulation kinetic of the free sugars and the ferulic acid esters. A correlation could be observed between the occurrence of the esters and sucrose in the pollentapetum fraction during the anther development.