Biosynthesis of mycothiol: elucidation of the sequence of steps in Mycobacterium smegmatis.

Several members of the Actinomycetales, including the medically important mycobacteria, produce 1-D-myo-inosityl-2-(N-acetyl-L-cysteinyl)amino-2-deoxy-alpha-D- glucop yranoside (trivial name mycothiol) as their principal low-molecular-mass thiol. The pseudo-disaccharide component of mycothiol, 1-D-myo-inosityl-2-amino-2-deoxy-alpha-D-glucopyranoside (alpha-D-GI), was synthesized by ligation of 1-D,L-2,3,4,5, 6-penta-O-acetyl-myo-inositol to 3,4,6-tri-O-acetyl-2-deoxy- 2-(2,4-dinitrophenylamino)-alpha-D-glu- copyranosyl bromide to give, in the first instance, an isomeric mixture of alpha- and beta-linked pseudo-disaccharides. The alpha-coupled D,D and D,L isomers, alpha-D-GI and alpha-L-GI respectively, were purified from the mixture by TLC, followed by removal of the protecting groups. A cell-free extract of Mycobacterium smegmatis catalysed the ligation of cysteine, acetate and alpha-D-GI in the presence of ATP and Mg2+ to form mycothiol, as judged by HPLC. When no acetate was added to the incubation mixture, an additional thiol accumulated. In the presence of [14C]acetate no radiolabel was recovered in this species, but only in mycothiol. The additional thiol was isolated as the bimane derivative, and 1H and 1H-1H COSY NMR spectra confirmed its identity as desacetylmycothiol. A more complete conversion of desacetylmycothiol into mycothiol was achieved in the presence of acetyl-S-CoA. These results indicate that the biosynthesis of mycothiol proceeds by the sequential addition of cysteine and acetate to alpha-D-GI. The inositol moiety appears to be an important determinant of specificity, since alpha-L-GI was poorly utilized.

Physiological characteristics of rugby players including muscle glycogen content and muscle fibre composition.

There are few studies of the anthropomorphic and physiological characteristics of South African rugby players. Twenty-nine club rugby players were evaluated for body composition, maximal treadmill performance, muscle fibre composition and the muscle glycogen content, including the effects of carbohydrate-loading and playing a rugby match. The body composition and physiological characteristics of the players were similar to that previously reported for international players. Very high absolute values for maximum oxygen consumption (VO2 max) were measured in the forwards. Both backs and forwards had a preponderance of fast-twitch muscle fibres (57% and 53% respectively). Carbohydrate-loading increased pre-match muscle glycogen content but was unnecessary since severe muscle glycogen depletion did not occur during a match even in players on a normal mixed diet before competition. It is concluded that for forwards, a high absolute VO2 max, and for both backs and forwards a predominance of fast-twitch muscle fibres are pre-requisites for success in this sport. Pre-match carbohydrate-loading would appear to be necessary only when more than one match is to be played within 48 hours.