Sugar Utilization and Its Control in Hyperthermophiles
Willem M. DE VOS*
Department of Biomolecular Sciences, Wageningen Agricultural University, The Netherlands
Many hyperthermophilic microorganisms show rapid heterotrophic growth on a variety of carbohydrates, including hexoses, pentoses, and polymeric sugars. There has been considerable fundamental and applied interest in the utilization of glucose and its α- and β- linked polymers by hyperthermophilic Archaea differs from the canonical glycolytic pathways, involves novel enzymes, and shows a unique control. This will be illustrated by recent physiological, biochemical and genetic work on Pyrococcusfuriosus, which is capable of fermenting the marine polymer laminarin, a β-1,3-1inked polymer of glucose.
The first steps in the degradation of laminarin include its depolymerization by a secreted endoglucanase (LamA) and the intracellular hydrolysis of the resulting oligosaccharides by a β-glycosidase (CelB) into glucose. Both enzymes have been functionally overproduced in E. coli allowing their detailed biochemical characterization, protein engineering, and crystallization trials. Genetic studies have shown that the lamA and celB genes are located in a divergent operon, the expression of which is controlled by the extracellular sugar concentration. Transcription studies of the intergenic region revealed the presence of two functional promoters that showed a back-to-back configuration. The initiation of transcription was found to be induced by growth on β-1inked sugars, most likely via transcription activation.
The resulting glucose is utilized by a modified Embden-Meyerhof pathway involving at least three novel enzymes, two ADP-dependent kinases - glucokinase and phospho-fructokinase - and a glyceraldehyde-3-phosphate:ferredoxin oxidoreducase (Gor). The gor gene has been characterized and found to encode a [4Fe-4S] protein with significant similarity to the structurally well-studied aldehyde:ferredoxin oxidoreductase from P. furiosus, including conserved binding sites for a W-dipterin that is likely to be involved in catalysis. Further studies showed that the transcription of the gor gene is induced during glycolysis. Moreover, the activity of the glyceraldehyde-3-P dehydrogenase, which appears to be exclusively involved in gluconeogenesis, is repressed under glycolytic conditions. This suggests that the conversion of glyceraldehyde-3-P to 3-P-glycerate may be a novel site of glycolytic regulation. The distribution of Gor-like proteins, their possible function, and their control in other microorganisms will be discussed.
