Nucleic Acids Research Advance Access originally published online on December 20, 2007
Nucleic Acids Research 2008 36(4):1072-1080; doi:10.1093/nar/gkm1121
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Nucleic Acids Research, 2008, Vol. 36, No. 4 1072-1080
© 2007 The Author(s)
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.0/uk/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
RNA |
Kluyveromyces lactis 
-toxin, a ribonuclease that recognizes the anticodon stem loop of tRNA
Department of Molecular Biology, Umeå University, 901 87 Umeå, Sweden
*To whom correspondence should be addressed. Tel: +46 90 7856764; Fax: +46 90 772630; Email: anders.bystrom{at}molbiol.umu.se
Received October 6, 2007. Revised November 30, 2007. Accepted December 1, 2007.
Kluyveromyces lactis 
-toxin is a tRNA endonuclease that cleaves Saccharomyces cerevisiae 
, 
and 
between position 34 and position 35. All three substrate tRNAs carry a 5-methoxycarbonylmethyl-2-thiouridine (mcm5s2U) residue at position 34 (wobble position) of which the mcm5 group is required for efficient cleavage. However, the different cleavage efficiencies of mcm5s2U34-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 -toxin. A synthetic minihelix RNA corresponding to the anticodon stem loop (ASL) of the natural substrate is cleaved at the same position as the natural substrate. In 
, the nucleotides U34U35C36A37C38 are required for optimal -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 
reinforced the strong stimulatory effects of the mcm5 group, revealed a weak positive effect of the s2 group and a negative effect of the bacterial 5-methylaminomethyl (mnm5) group. The data underscore the high specificity of this yeast tRNA toxin.
Present address: Jian Lu, Laboratory of Molecular Gerontology, National Institute on Aging, 5600 Nathan Shock Drive, Baltimore, MD 21224-6825, USA
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