C.cylindracea tRNA, none of the mutations at position 33 caused leucylation of the tRNA, as was observed with the native tRNA Ser CAG, and there was no reduction in of G at position 33, where a pyrimidine (mostly U) is completely conserved in usual tRNAs (Sprinzl et al., serylation activity ( Figure 3C ). In contrast, the replacement of G33 by pyrimidines in C.zeylanoides tRNA Ser CAG 1996). Since we considered it is possible that this notable feature may be in some way related to the unusual considerably enhanced the leucylation activity ( Figure 3B , right-hand graph), while no significant difference was aminoacylation characteristics described above and/or to the translation of non-universal genetic code, we examined observed in the serylation activity ( Figure 3B , left-hand graph). The kinetic parameters of leucylation for the the effect of residue 33 on the aminoacylation and transla- show the spots corresponding to acetylleucine and acetylserine as markers, respectively. (D) Analysis of acetylamino acids attached to tRNA fragments on a TLC plate. Lane 2 shows the spot corresponding to the acetylamino acids derived from the RNase T1 fragment of C.zeylanoides tRNA Ser CAG. Lanes 1 and 3 indicate the spots corresponding to acetylleucine and acetylserine, respectively. Ten micrograms of [ 14 C]acetylaminoacyl-tRNA Ser CAG from C.zeylanoides was digested with RNase T1 and developed on cellulose TLC plates under the same conditions as (C). CCA fragments with [ 14 C]acetylamino acids were scraped from the plate from which the fragments were eluted with H20 and desalted by Sep-pak C18 under the conditions described in the literature (Wang et al., 1990) .