Guanosine Diphosphate-4-keto-d-rhamnose Reductase

Abstract

Abstract The pyridine nucleotide-mediated reduction of GDP-4-keto-6-deoxy-d-mannose (GDP-4-keto-d-rhamnose) as catalyzed by extracts of soil bacterium GS (ATCC 19241) has been investigated with the use of 200- to 400-fold purified GDP-4-keto-d-rhamnose reductase. The results of these investigations suggest that a single enzyme protein is responsible for the formation of both C-4 epimers, GDP-d-rhamnose and GDP-d-talomethylose. Experiments with GDP-d-[14C]rhamnose exclude interconversion of the sugar nucleotide epimers by means of a GDP-d-rhamnose-4-epimerase. Thus, their formation appears to result from separate reductive reactions in which either GDP-d-rhamnose or GDP-d-talomethylose is produced, depending upon the orientation of the pyranose ring with respect to the reducing hydrogen atom. In experiments with stereospecifically labeled [4-3H]TPNH, it has been possible to show the quantitative transfer of tritium from [4B-3H]TPNH to the rhamnose and talomethylose moieties of GDP-d-rhamnose and GDP-d-talomethylose. Hence, while location of tritium at C-4 of the 6-deoxyhexose is assumed, it appears that a single enzyme transfers hydrogen from the 4B position of TPNH to either side of the pyranose ring at the C-4 carbonyl function of GDP-4-keto-d-rhamnose. This is the first description of a pyridine nucleotide oxidoreductase which lacks precise stereoselectivity with respect to its hydrogen receptor carbonyl function.