Figure 3.

Characterization of the T.maritima RNase P holoenzyme. Initial velocities of cleavage of precursor tRNA^Tyr were determined as follows: 1 nM RNA and 10 nM protein were preincubated in their respective reaction buffers for 5 min at 50°C (see text). The reactions were started by the addition of 100 nM ³²P-labeled substrate. Aliquots were taken at 1 min intervals and separated on 8% polyacrylamide/7 M urea denaturing gels. The amount of cleavage was determined using a phosphoimager, and is indicated as nmol product/nmol RNase P RNA (min). Thermotoga maritima RNase P holoenzyme dependence on (A) KCl (in 50 mM Tris, pH 7.5, 10 mM MgCl₂ in this and other cases that follow), (B) NaCl, (C) NH₄Cl, (D) MgCl₂ (in 50 mM Tris, pH 7.5, 400 mM NH₄Cl).