LYOTROPIC-SALT-INDUCED CHANGES IN MONOMER DIMER/TETRAMER ASSOCIATION EQUILIBRIUM OF FORMYLTRANSFERASE FROM THE HYPERTHERMOPHILIC METHANOPYRUS-KANDLERI IN RELATION TO THE ACTIVITY AND THERMOSTABILITY OF THE ENZYME/

Formyltransferase from Methanopyrus kandleri is composed of only one t ype of subunits of molecular mass 32 kDa. The enzyme requires the pres ence of lyotropic salts for activity and thermostability. We report he re that the enzyme is in a monomer/dimer/tetramer association equilibr ium, the association constant being affected by lyotropic salts. At 0. 01 M K2HPO4/KH2PO4, pH 7.2, the enzyme (0.4 mg/ml) was mainly present in a monomeric form. Upon increase of the phosphate concentration, the concentration of the dimer increased up to a phosphate concentration of 0.6 M, then decrease at the expense of tetramer formation up to a p hosphate concentration of 1.0 M. The specific activity at 4 degrees C increased from <0.1 U/mg at 0.01 M, over 1.5 U/mg at 0.6 M to 3.6 U/mg at 1.0 M. Similar results were obtained with ammonium sulfate as lyot ropic salt. The findings indicate that both oligomerization and activi ty increase with increasing salt concentrations, suggesting that there is a causal connection. To determine this, we exploited the observati on that oligomer formation was not induced by the weak lyotropic salt NaCl up to a concentration of 1.5 M and that the dissociation of the d imer into the monomer at 4 degrees C proceeded very slowly (50% in app roximately 6 h). This allowed us to study the effect of NaCl on the ac tivity of the oligomers at NaCl concentrations not sufficient to induc e oligomerization. At 4 degrees C, the activity of the oligomers incre ased from 0.3 U/mg at 0.25 M NaCl to 3.4 U/mg at 1.0 M NaCl. At these NaCl concentrations, the monomers were inactive. The findings indicate that oligomerization is a prerequisite for enzyme activity in the pre sence of NaCl. The salt-dependent induction of oligomerization was par allelled by an increase in thermostability; strong lyotropic salts con ferred thermostability at much lower concentrations than the weak lyot ropic NaCl.