Studies on trehalose phosphate synthetase in Mycobacterium smegmatis
Liu, Cecillia Huan
Elbein, Alan D.
Master of Arts
The biosynthesis of trehalose phosphate occurs by two reactions which utilize different sugar nucleotides, UDP-glucose or GDP-glucose. Cell free extracts of several mycobacteria catalyzed the synthesis of trehalose phosphate from both UDPglucose and GDP-glucose. An extract of Mycobacterium smegmatis was separated into two fractions by chromatography on DEAE cellulose. Fraction 1 catalyzed the synthesis of trehalose (trehalose-P) from GDP-glucose but only slightly active with UDP-glucose. However, when fraction 2 was added to fraction 1, UDP-glucose was able to serve as an effective glucosyl donor for trehalose synthesis. Under these conditions, GDP-glucose was still active. Fraction 2 had no detectable enzymatic activity and was heat-stable. The action of fraction 2 could be replaced by crude a -lactalbumin, but not by pure a -lactalbumin, lysozyme, albumin or a number of other proteins. The products formed from UDP-glucose-14C and fractions 1 and 2 or GDP-glucose-14C and fraction' 1 were identified as trehalose by paper chromatography and cocrystallization of the octaacetate derivative to constant specific activity with authentic trehalose octaacetate. As to the substrate specificity, ADP-glucose gave similar results to UDP-glucose. Fructose-6-P was equally effective in fulfilling the requirement for glucose-6-P, presumably due to the presence of phosphohexose isomerase.