Source:http://linkedlifedata.com/resource/pubmed/id/10828971
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rdf:type | |
lifeskim:mentions | |
pubmed:issue |
21
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pubmed:dateCreated |
2000-7-11
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pubmed:abstractText |
Escherichia coli dimethylallyl diphosphate:tRNA dimethylallyltransferase (DMAPP-tRNA transferase) catalyzes the alkylation of the exocyclic amine of A37 by a dimethylallyl unit in tRNAs with an adenosine in the third anticodon position (position 36). By use of purified recombinant enzyme, steady- state kinetic studies were conducted with chemically synthesized RNA oligoribonucleotides to determine the essential elements within the tRNA anticodon stem-loop structure required for recognition by the enzyme. A 17-base oligoribonucleotide corresponding to the anticodon stem-loop of E. coli tRNA(Phe) formed a stem-loop minihelix (minihelix(Phe)) when annealed rapidly on ice, while the same molecule formed a duplex structure with a central loop when annealed slowly at higher concentrations. Both the minihelix and duplex structures gave k(cat)s similar to that for the normal substrate (full-length tRNA(Phe) unmodified at A37), although the K(m) for minihelix(Phe) was approximately 180-fold higher than full-length tRNA. The A36-A37-A38 motif, which is completely conserved in tRNAs modified by the enzyme, was found to be important for modification. Changing A36 to G in the minihelix resulted in a 260-fold reduction in k(cat) compared to minihelix(Phe) and a 13-fold increase in K(m). An A38G variant was modified with a 9-fold reduction in k(cat) and a 5-fold increase in K(m). A random coil 17-base oligoribonucleotide in which the loop sequence of E. coli tRNA(Phe) was preserved, but the 5 base pair helix stem was completely disrupted and showed no measurable activity, indicating that a helix-loop structure is essential for recognition. Finally, altering the identity of several base pairs in the helical stem did not have a major effect on catalytic efficiency, suggesting that the enzyme does not make base-specific contacts important for binding or catalysis in this region.
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pubmed:grant | |
pubmed:language |
eng
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pubmed:journal | |
pubmed:citationSubset |
IM
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pubmed:chemical |
http://linkedlifedata.com/resource/pubmed/chemical/Alkyl and Aryl Transferases,
http://linkedlifedata.com/resource/pubmed/chemical/Anticodon,
http://linkedlifedata.com/resource/pubmed/chemical/Oligoribonucleotides,
http://linkedlifedata.com/resource/pubmed/chemical/RNA, Bacterial,
http://linkedlifedata.com/resource/pubmed/chemical/RNA, Transfer,
http://linkedlifedata.com/resource/pubmed/chemical/adenylate isopentenyltransferase
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pubmed:status |
MEDLINE
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pubmed:month |
May
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pubmed:issn |
0006-2960
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pubmed:author | |
pubmed:issnType |
Print
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pubmed:day |
30
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pubmed:volume |
39
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pubmed:owner |
NLM
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pubmed:authorsComplete |
Y
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pubmed:pagination |
6546-53
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pubmed:dateRevised |
2007-11-14
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pubmed:meshHeading |
pubmed-meshheading:10828971-Alkyl and Aryl Transferases,
pubmed-meshheading:10828971-Anticodon,
pubmed-meshheading:10828971-Base Sequence,
pubmed-meshheading:10828971-Escherichia coli,
pubmed-meshheading:10828971-Kinetics,
pubmed-meshheading:10828971-Nucleic Acid Conformation,
pubmed-meshheading:10828971-Nucleic Acid Denaturation,
pubmed-meshheading:10828971-Oligoribonucleotides,
pubmed-meshheading:10828971-RNA, Bacterial,
pubmed-meshheading:10828971-RNA, Transfer,
pubmed-meshheading:10828971-Substrate Specificity
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pubmed:year |
2000
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pubmed:articleTitle |
Escherichia coli dimethylallyl diphosphate:tRNA dimethylallyltransferase: essential elements for recognition of tRNA substrates within the anticodon stem-loop.
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pubmed:affiliation |
Department of Chemistry, University of Utah, Salt Lake City 84112, USA.
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pubmed:publicationType |
Journal Article,
Research Support, U.S. Gov't, P.H.S.
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