Molecular basis and its application of formaldehyde-fixing reactions in bacteria and Archaea.

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Hexulose-6-phosphate synthase(Hps) and Phosphohexuloisomerase(Phi), two key enzymes in the ribulose monophosphate pathway, has been recently found in a number of bacterial and archaeal strains. In this study, we studied on (i)the regulation of the expresseion of hps and phi gene in bacteria, and (ii)enzymological properties and their physiological roles of hps and phi in archaea.(i)Two regulatory proteins (RmpR from a methylotroph Mycobacterium gastri MB19 and HxlR from a non-methylotroph Bacillus subtilis) were produced in E.coli and their abilities to DNA were tested. HxlR bound to the upstream region of the hps/phi operon and was shown to act as a positive regulator of the formaldehyde-inducible gene expression.(ii)The hyperthermophilic archaeon Pyrococcus horikoshii had a fused protein consisting of Hps and Phi. The entire fusion protein and separate Hps and Phi polypeptides were produced in E.coli, purified and biochemically characterized. The fused protein was found in a particulate fraction. It was suggested that the thermostability of the Phi moiety of the fused protein resulted from fusion with Hps. Fusion of Hps and Phi increased the catalytic efficiency of the net reaction. On the other hand the methanogenic archaeon Methanosarcina mazei Goel had a fused protein consisting of Hps and formaldehyde activating enzyme (Fae). Hps-Fae fused protein produced in E.coli exhibited both Hps and Phi activities. From the catalytic properties of Hps-Phi and Hps-Fae proteins from two archeal strains, it was suggested that these proteins are involved not only in the formaldehyde fixation (forward reaction) but also in the generation of ribose 5-phosphate (reverse reaction).