THE BIOSYNTHESIS PATHWAY OF DI-MYO-INOSITOL-1,1'-PHOSPHATE IN PYROCOCCUS-WOESEI

From in vitro experiments with dialyzed cell-free extracts of the hype rthermophilic Archaeum Pyrococcus woesei, the biosynthetic pathway of di-myo-inositol-1,1'-phosphate was deduced. Starting from glucose-6-ph osphate, the synthesis proceeds in two steps with L-myo-inositol 1-pho sphate as intermediate. (1) Interconversion of glucose 6-phosphate to inositol l-phosphate was observed without adding cofactors, such as nu cleoside triphosphates or pyridine dinucleotides suggesting that the f irst enzyme reaction corresponds to the NAD(+)-dependent inositol 1-ph osphate synthase reaction as already described for eukaryal systems, b ut differing from the latter by a stronger pyridine dinucleotide bindi ng rendering the enzyme virtually independent from external NAD(+). (2 ) In a second step, two L-myo-inositol 1-phosphates are coupled under the expense of NTP to yield di-myo-inositol-1,1'-phosphate. The coupli ng of two L-myo-inositol 1-phosphates without preceding dephosphorylat ion of one of both by a phosphorylase as proposed for the di-myo-inosi tol-1,1'-phosphate synthesis in Methanococcus igneus (Chen, L. and Rob erts, M. (1998) Appl. Environ. Microbiol. 64, 2609-2615) is supported by labeling experiments which resulted only in a labeled product with L-myo [U-14C]inositol 1-phosphate, but not with radiolabeled L-myo-[U- C-14]inositol and non-labeled L-myo-inositol 1-phosphate. (C) 1998 Fed eration of European Microbiological Societies. Published by Elsevier S cience B.V. All rights reserved.