Figure 4.

The Mn²⁺-rescue assays for T.maritima tRNase Z variants containing single substitutions at Asp-52 or His-222. (A) Protein profiles of histidine-tagged T.maritima tRNase Z variants. Each purified protein (0.5−1 μg) was analyzed on an SDS–10% polyacrylamide gel, and visualized by staining the gel with Coomassie brilliant blue R-250. (B) The assays for in vitro 3′ processing of T.maritima pre-tRNA^Arg(₇₄CCA₇₆) by T.maritima tRNase Z variants. The pre-tRNA (0.1 pmol) was incubated with the tRNase Z variants (4 pmol) at 60°C for 10 min. The cleavage products were analyzed on a 10% polyacrylamide–8 M urea gel.