Kluyveromyces lactis gamma-toxin, a ribonuclease that recognizes the anticodon stem loop of tRNA.
Kluyveromyces lactis gamma-toxin is a tRNA endonuclease that cleaves Saccharomyces cerevisiae [see text] between position 34 and position 35. All three substrate tRNAs carry a 5-methoxycarbonylmethyl-2-thiouridine (mcm(5)s(2)U) residue at position 34 (wobble position) of which the mcm(5) group is required for efficient cleavage. However, the different cleavage efficiencies of mcm(5)s(2)U(34)-containing tRNAs suggest that additional features of these tRNAs affect cleavage. In the present study, we show that a stable anticodon stem and the anticodon loop are the minimal requirements for cleavage by gamma-toxin. A synthetic minihelix RNA corresponding to the anticodon stem loop (ASL) of the natural substrate [see text] is cleaved at the same position as the natural substrate. In [see text], the nucleotides U(34)U(35)C(36)A(37)C(38) are required for optimal gamma-toxin cleavage, whereas a purine at position 32 or a G in position 33 dramatically reduces the cleavage of the ASL. Comparing modified and partially modified forms of E. coli and yeast [see text] reinforced the strong stimulatory effects of the mcm(5) group, revealed a weak positive effect of the s(2) group and a negative effect of the bacterial 5-methylaminomethyl (mnm(5)) group. The data underscore the high specificity of this yeast tRNA toxin.
Lu
Jian
J
Department of Molecular Biology, Umeå University, 901 87 Umeå, Sweden.
Esberg
Anders
A
Huang
Bo
B
Byström
Anders S
AS
D000926:Anticodon / Q000737:chemistry* / Q000378:metabolism; D001483:Base Sequence; D004722:Endoribonucleases / Q000378:metabolism*; D007716:Kluyveromyces / Q000201:enzymology*; D008969:Molecular Sequence Data; D009154:Mutation; D009183:Mycotoxins / Q000378:metabolism*; D012351:RNA, Transfer, Gln / Q000737:chemistry / Q000378:metabolism; D012352:RNA, Transfer, Glu / Q000737:chemistry / Q000378:metabolism; D012357:RNA, Transfer, Lys / Q000737:chemistry / Q000378:metabolism; D013379:Substrate Specificity; D013891:Thiouridine / Q000031:analogs & derivatives / Q000737:chemistry
D000926:Anticodon; D009183:Mycotoxins; D012351:RNA, Transfer, Gln; D012352:RNA, Transfer, Glu; D012357:RNA, Transfer, Lys; D013891:Thiouridine; C011916:5-methylaminomethyl-2-thiouridine; D004722:Endoribonucleases