FORMATION OF GUANOSINE DIPHOSPHATE FUCOSE FROM


Enzymatic Evidence for the Configuration of d-Isocitric Acid. ALTON MEISTER , MARYELLEN STRASSBURGER. Nature 1963 200 (4903), 259-260 ...
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Iiuried in the electron density of the cobalt atom as has been suggested' previously whereas these numerically accurate calculations provide no support for a Co-H distance of -2.0. DEPARTMEST O F CHEMISTRY MASSACHUSETTS INSTITUTE O F CANBRIDGE 39, MASS

TECHNOLOGY F.A.

RECEIVED JULY

COTTON

7, 1958

FORMATION O F GUANOSINE DIPHOSPHATE FUCOSE FROM GUANOSINE DIPHOSPHATE MANNOSE

Sir: A fucose containing nucleotide, guanosine diphosphate fucose, has been isolated recently from sheep milk' and from Aerobacter aerogenes.2 I t has now been found that GDPM3can be converted to guanosine diphosphate fucose by dialysed crude extracts of A . aerogenes4 in the presence of TPNH. The conversion was detected as follows: GDPAI, prepared from yeast,5 was incubated with T P N H and crude bacterial extracts obtained by shaking the cells with glass beads6 followed by centrifugation and dialysis. The guanosine sugar nucleotides were then isolated and purified from the incubation mixtures by charcoal adsorption and paper ~ h r o m a t o g r a p h y . ~Chromatography of the sugar liherated by 0.01 N HCl hydrolysis of the isolated nucleotides revealed, in addition to mannose, the presence of a second compound. This new compound exhibited the characteristic 400 mp absorption peak when examined by the specific colorimetric assay for 6-deoxyhexose.' Upon paper chromatography, the unknown sugar co-chromatographed with authentic fucose using the solvents 2TABLE I COYVERSIONOF GUANOSINE;DIPHOSPHATEMANNOSE TO GUANOSISF: DIPHOSPHATE FUCOSE The reaction mixtures contained 1.0 pmole G D P M , 3 mg. of crude extract protein and additions in 1.0 ml. of 0.05 M tris-(Ii?-droxj~meth~I)-aminomethane buffer, p H 7.8. Incuhation was carried out a t 37' for 4 hours. T h e nucleotides were then adsorbed on charcoal and the nucleotide 1)nunrl sugars liherated by heating for ten minutes a t 100" i n 0.01 .V H C 1 . Xfter deionization with Amberlite MR-3, tlic fucose in the hydrolysate was estimated colorimetrically7 n r b y paper chromatography. Additions

&mole fucose formed