The effects of spermine and Mg2+ on the catalytic mechanism of isoleucine: tRNA ligase.
Isoleucyl-tRNA formation catalysed by isoleucine: tRNA ligase is stimulated by both Mg2+ and spermine in the pH-range 7.0 to 8.0 at 310 K. At low [Mg2+] the acceleration caused by both cations together exceeds the sum of their individual effects. 2. The spermine-stimulated reaction has a steeper temperature-dependence than reaction in the presence of Mg2+. Two phases in the kinetics of isoleucyl-tRNA formation are detected in the presence of Mg2+ plus or minus spermine, but only a single step is observed in the presence of spermine alone. Thus the rate-limiting steps under normal assay conditions are different for the two cations. 3. Enzyme-bound isoleucyl-AMP can be formed in the absence of Mg-2+ and plus or minus spermine. 4. It is concluded that there is no evidence for cation-dependent differences in the reaction mechanism of isoleucine: tRNA ligase, though there are certainly differences in the relative rates of some of the individual steps.
D004789:Enzyme Activation; D004926:Escherichia coli / Q000201:enzymology*; D006863:Hydrogen-Ion Concentration; D007532:Isoleucine; D007700:Kinetics; D008025:Ligases; D008274:Magnesium; D012343:RNA, Transfer; D013096:Spermine; D013696:Temperature
D007532:Isoleucine; D013096:Spermine; D012343:RNA, Transfer; D008025:Ligases; D008274:Magnesium