Studies of Sulfate Utilization by Algae. 4. Properties of a Cell-Free Sulfate-Reducing System From Chlorella

Abstract

A cell-free system from Chlorella pyrenoidosa (Emerson strain 3) which produces acid-volatile radioactivity from SO3542- is described. A high speed supernatant from cells broken in the French Press at pH 7.0 shows maximal activity when fortified with ATP, an ATP-generating system (creatine phosphate and creatine phosphoMnase), TPN, a TPN-reducing system (glucose-6-phosphate and glucose-6-phosphate dehy-drogenase) and MgCl2. This system is quite labile and is not stable to dialysis. Additionof low concentrations of 2,3,-dimercaptopropan-l-ol (HAL) to the buffers used for enzyme preparation stabilize the extracts and permit them to be dialyzed for 4 hours without loss of activity. If additional BAL is also added to the incubation mixtures it can replace TPNH as a reductant. DPNH also shows appreciable activity. The system prepared with BAL-containing buffers shows maximal activity at pH 9.0. At this pH, the system requires only ATP, Mg2+ and additional BAL and has high activity and stability compared with the other conditions tried. The optimum concentrations of these reactants has been determined and the kinetics of production of acid-volatile radioactivity are described. Nucleoside triphosphates other than ATP are not appreciably active in this system. In all cases, anaerobic conditions are required for maximal activity, the enzyme extracts are labile to heat, and no unequivocal requirement for thioctic acid can be demonstrated.