Discrimination of tRNA(Leu) isoacceptors by the insertion mutant of Escherichia coli leucyl-tRNA synthetase

Tong Li; Li Yong; Ni Ni Guo; En Duo Wang; Ying Lai Wang

doi:10.1021/bi9901984

Discrimination of tRNA(Leu) isoacceptors by the insertion mutant of Escherichia coli leucyl-tRNA synthetase

Tong Li, Li Yong, Ni Ni Guo, En Duo Wang, Ying Lai Wang

Research output: Contribution to journal › Article › peer-review

17 Scopus citations

Abstract

A variant (LeuRS-A) of Escherichia coli leucyl-tRNA synthetase (LeuRS) carrying a 40-residue duplication in its connective peptide 1 (CP1) has a 3- fold lower specificity for tRNA₁/(Leu) than for tRNA₂/(Leu), whereas wild- type LeuRS has the same specificity for these two isoacceptors. The replacement of the acceptor stem of tRNA₂/(Leu) with tRNA₁/(Leu) yields a chimeric tRNA(Leu) for which wild-type LeuRS has the same specificity as it does for the two normal isoacceptors mentioned, but for which LeuRS-A has a reduced specificity similar to that for tRNA₁/(Leu), indicating a difference between these two acceptor stems. LeuRS-A is slightly less stable than the native enzyme. Wild-type LeuRS and LeuRS-A have almost same K(d) value for their interaction with tRNA₁/(Leu) as determined by fluorescence quenching. No difference was detected between these two proteins by CD and fluorescence spectroscopy. These results show that LeuRS-A can discriminate between the two isoacceptors of tRNA(Leu).

Original language	English (US)
Pages (from-to)	9084-9088
Number of pages	5
Journal	Biochemistry
Volume	38
Issue number	28
DOIs	https://doi.org/10.1021/bi9901984
State	Published - Jul 13 1999
Externally published	Yes

ASJC Scopus subject areas

Biochemistry

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title = "Discrimination of tRNA(Leu) isoacceptors by the insertion mutant of Escherichia coli leucyl-tRNA synthetase",

abstract = "A variant (LeuRS-A) of Escherichia coli leucyl-tRNA synthetase (LeuRS) carrying a 40-residue duplication in its connective peptide 1 (CP1) has a 3- fold lower specificity for tRNA1/(Leu) than for tRNA2/(Leu), whereas wild- type LeuRS has the same specificity for these two isoacceptors. The replacement of the acceptor stem of tRNA2/(Leu) with tRNA1/(Leu) yields a chimeric tRNA(Leu) for which wild-type LeuRS has the same specificity as it does for the two normal isoacceptors mentioned, but for which LeuRS-A has a reduced specificity similar to that for tRNA1/(Leu), indicating a difference between these two acceptor stems. LeuRS-A is slightly less stable than the native enzyme. Wild-type LeuRS and LeuRS-A have almost same K(d) value for their interaction with tRNA1/(Leu) as determined by fluorescence quenching. No difference was detected between these two proteins by CD and fluorescence spectroscopy. These results show that LeuRS-A can discriminate between the two isoacceptors of tRNA(Leu).",

author = "Tong Li and Li Yong and Guo, {Ni Ni} and Wang, {En Duo} and Wang, {Ying Lai}",

year = "1999",

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T1 - Discrimination of tRNA(Leu) isoacceptors by the insertion mutant of Escherichia coli leucyl-tRNA synthetase

AU - Li, Tong

AU - Yong, Li

AU - Guo, Ni Ni

AU - Wang, En Duo

AU - Wang, Ying Lai

PY - 1999/7/13

Y1 - 1999/7/13

N2 - A variant (LeuRS-A) of Escherichia coli leucyl-tRNA synthetase (LeuRS) carrying a 40-residue duplication in its connective peptide 1 (CP1) has a 3- fold lower specificity for tRNA1/(Leu) than for tRNA2/(Leu), whereas wild- type LeuRS has the same specificity for these two isoacceptors. The replacement of the acceptor stem of tRNA2/(Leu) with tRNA1/(Leu) yields a chimeric tRNA(Leu) for which wild-type LeuRS has the same specificity as it does for the two normal isoacceptors mentioned, but for which LeuRS-A has a reduced specificity similar to that for tRNA1/(Leu), indicating a difference between these two acceptor stems. LeuRS-A is slightly less stable than the native enzyme. Wild-type LeuRS and LeuRS-A have almost same K(d) value for their interaction with tRNA1/(Leu) as determined by fluorescence quenching. No difference was detected between these two proteins by CD and fluorescence spectroscopy. These results show that LeuRS-A can discriminate between the two isoacceptors of tRNA(Leu).

AB - A variant (LeuRS-A) of Escherichia coli leucyl-tRNA synthetase (LeuRS) carrying a 40-residue duplication in its connective peptide 1 (CP1) has a 3- fold lower specificity for tRNA1/(Leu) than for tRNA2/(Leu), whereas wild- type LeuRS has the same specificity for these two isoacceptors. The replacement of the acceptor stem of tRNA2/(Leu) with tRNA1/(Leu) yields a chimeric tRNA(Leu) for which wild-type LeuRS has the same specificity as it does for the two normal isoacceptors mentioned, but for which LeuRS-A has a reduced specificity similar to that for tRNA1/(Leu), indicating a difference between these two acceptor stems. LeuRS-A is slightly less stable than the native enzyme. Wild-type LeuRS and LeuRS-A have almost same K(d) value for their interaction with tRNA1/(Leu) as determined by fluorescence quenching. No difference was detected between these two proteins by CD and fluorescence spectroscopy. These results show that LeuRS-A can discriminate between the two isoacceptors of tRNA(Leu).

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Discrimination of tRNA(Leu) isoacceptors by the insertion mutant of Escherichia coli leucyl-tRNA synthetase

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