The human homolog of Escherichia coli orn degrades small single-stranded RNA and DNA oligomers

被引:40
作者
Nguyen, LH [1 ]
Erzberger, JP [1 ]
Root, J [1 ]
Wilson, DM [1 ]
机构
[1] Univ Calif Lawrence Livermore Natl Lab, Mol & Struct Biol Div, Livermore, CA 94551 USA
关键词
D O I
10.1074/jbc.M002672200
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
We report here the identification of human homologues to the essential Escherichia coli Orn protein and the related yeast mitochondrial DNA-escape pathway regulatory factor Ynt20, The human proteins appear to arise from alternatively spliced transcripts, and are thus identical, except the human Ynt20 equivalent contains an NH2-terminal extension that possesses a predicted mitochondrial protease cleavage signal. In vitro analysis revealed that the smaller human protein exhibits a 3' to 5' exonuclease activity for small (primarily less than or equal to 5 nucleotides in length) single-stranded RNA and DNA oligomers, We have named this human protein Sfn for Small fragment nuclease to reflect its broad substrate range, and have termed the longer protein hSfn alpha. Sfn prefers Mn2+ as a metal cofactor and displays a temperature-resistant (to 50 degrees C) nuclease activity. Kinetic analysis indicates that Sfn exhibits a similar affinity for small RNAs and DNAs (K-m of similar to 1.5 mu M), but degrades small RNAs similar to 4-fold more efficiently than DNA, Mutation of a conserved aspartate (Asp(136)) to alanine abolishes both nuclease activities of Sfn, Northern blot analysis revealed that a l-kilobase transcript corresponding to SFN and/or SFN alpha (these mRNAs differ by only two nucleotides) is expressed at varying levels in all fetal and adult human tissues examined. Expressed tag sequence clone analysis found that the two splice variants, SFN to SFN alpha, are present at a ratio of roughly 4 to 1, respectively. The results presented within suggest a role for human Sfn in cellular nucleotide recycling.
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页码:25900 / 25906
页数:7
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