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PMID: 8233780 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S.

Contributions of discrete tRNA(Ser) domains to aminoacylation by E.coli seryl-tRNA synthetase: a kinetic analysis using model RNA substrates.

Nucleic acids research ·Vol. 21 ·No. 19 ·1993-09-25 ·Pages 4467-75

Sampson JR, Saks ME

Abstract

The aminoacylation kinetics of T7 transcripts representing defined regions of Escherichia coli serine tRNAs were determined using purified E.coli seryl-tRNA synthetase (SerRS) and the kinetic values were used to estimate the relative contribution of various tRNA(Ser) domains to recognition by SerRS. The analysis revealed that the extra stem/loop structure, characteristic of type II tRNAs such as tRNA(Ser), is the domain which makes the largest contribution to kcat/Km of aminoacylation. Moreover, Km of aminoacylation was increased by a factor of about 1000 when the extra stem/loop was changed to the consensus sequence of type I tRNA extra loops indicating that the stem structure contributes significantly to the binding of tRNA(Ser) to SerRS. A model RNA, which represents only the tRNA(Ser) coaxial acceptor-T psi C stem/loop domain, was also specifically aminoacylated by SerRS having a kcat/Km about 1000-fold greater than background levels. A significant portion of the contribution of this domain to aminoacylation is attributable to the acceptor stem sequence making the acceptor stem the second most important domain for recognition by SerRS. Finally, kcat/Km was essentially unchanged when the entire anticodon stem/loop of tRNA(Ser) was deleted indicating that neither the anticodon nucleotides nor the surrounding stem/loop structure are important for recognition by SerRS.

MeSH Terms
Amino Acid Sequence Cloning, Molecular Escherichia coli/enzymology Kinetics Molecular Sequence Data Nucleic Acid Conformation RNA, Transfer, Ser/metabolism RNA-Binding Proteins/metabolism Serine-tRNA Ligase/metabolism Structure-Activity Relationship Transfer RNA Aminoacylation
Chemicals
RNA, Transfer, Ser RNA-Binding Proteins Serine-tRNA Ligase
Authors & Affiliations
2 authors, click to expand affiliations / ORCID
Sampson J R
Division of Biology 147-75, California Institute of Technology, Pasadena 91125.
Saks M E
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34 references, click to expand
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Article Info
Journal
Nucleic acids research
Abbr.
Nucleic Acids Res
ISSN
0305-1048
DOI
10.1093/nar/21.19.4467
Published
1993-09-25
Pages
4467-75
Language
English
Region
England
NLM ID
0411011
PMCID
PMC311177
Subset
IM
Grants
NIGMS NIH HHS · GM13776 · United States
NIGMS NIH HHS · GM48560 · United States
External Links
PubMed source DOI: 10.1093/nar/21.19.4467 PMC Full Text View journal details
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