Biomaterial Database

Reaction of modified and unmodified tRNA(Tyr) substrates with tyrosyl-tRNA synthetase (Bacillus stearothermophilus). 1993

J M Avis, and A G Day, and G A Garcia, and A R Fersht

Department of Chemistry, University of Cambridge, United Kingdom.

Three species of tRNA(Tyr) have been examined as substrates for the transfer reaction of the tyrosyl-tRNA synthetase (TyrRS) from Bacillus stearothermophilus: Escherichia coli tRNA(Tyr), B. stearothermophilus tRNA(Tyr) expressed in E. coli, and B. stearothermophilus tRNA(Tyr) that has been transcribed in vitro. The binding of the first two substrates to TyrRS may be readily monitored by stopped-flow studies of tryptophan fluorescence to give the rate and equilibrium constants. The in vitro-transcribed tRNA(Tyr), which lacks the modified bases queuosine and 2-(methylthio)-N6-isopentenyladenosine in the anticodon loop, does not cause a significant change in tryptophan fluorescence upon binding. The three tRNA(Tyr) substrates exhibit very similar steady-state kinetics in the charging reaction. Pre-steady-state kinetics of the transfer reaction, monitored by stopped-flow measurements of the change in protein fluorescence on the addition of tRNA(Tyr) to the E.Tyr-AMP complex, show two exponential changes for the modified tRNA(Tyr) substrates. The first is that due to substrate binding. The second has an identical rate to the single change observed for the reaction with the in vitro-transcribed tRNA(Tyr) and to that monitored by quenched-flow measurements on the formation of Tyr-tRNA(Tyr). Hence, the transfer reaction can be observed by stopped-flow. The dissociation constants (KtRNA) of tRNA from the enzyme and rates of tyrosine transfer (k4) show that all three tRNA molecules are kinetically equivalent substrates for TyrRS. The value of k4 is also similar to that found for authentic tRNA(Tyr) from B. stearothermophilus.(ABSTRACT TRUNCATED AT 250 WORDS)

UI	MeSH Term	Description	Entries
D007700	Kinetics	The rate dynamics in chemical or physical systems.
D004926	Escherichia coli	A species of gram-negative, facultatively anaerobic, rod-shaped bacteria (GRAM-NEGATIVE FACULTATIVELY ANAEROBIC RODS) commonly found in the lower part of the intestine of warm-blooded animals. It is usually nonpathogenic, but some strains are known to produce DIARRHEA and pyogenic infections. Pathogenic strains (virotypes) are classified by their specific pathogenic mechanisms such as toxins (ENTEROTOXIGENIC ESCHERICHIA COLI), etc.	Alkalescens-Dispar Group,Bacillus coli,Bacterium coli,Bacterium coli commune,Diffusely Adherent Escherichia coli,E coli,EAggEC,Enteroaggregative Escherichia coli,Enterococcus coli,Diffusely Adherent E. coli,Enteroaggregative E. coli,Enteroinvasive E. coli,Enteroinvasive Escherichia coli
D001411	Geobacillus stearothermophilus	A species of GRAM-POSITIVE ENDOSPORE-FORMING BACTERIA in the family BACILLACEAE, found in soil, hot springs, Arctic waters, ocean sediments, and spoiled food products.	Bacillus stearothermophilus,Bacillus thermoliquefaciens
D012365	RNA, Transfer, Tyr	A transfer RNA which is specific for carrying tyrosine to sites on the ribosomes in preparation for protein synthesis.	Transfer RNA, Tyr,Tyrosine-Specific tRNA,tRNATyr,tRNA(Tyr),RNA, Tyr Transfer,Tyr Transfer RNA,Tyrosine Specific tRNA,tRNA, Tyrosine-Specific
D013050	Spectrometry, Fluorescence	Measurement of the intensity and quality of fluorescence.	Fluorescence Spectrophotometry,Fluorescence Spectroscopy,Spectrofluorometry,Fluorescence Spectrometry,Spectrophotometry, Fluorescence,Spectroscopy, Fluorescence
D013816	Thermodynamics	A rigorously mathematical analysis of energy relationships (heat, work, temperature, and equilibrium). It describes systems whose states are determined by thermal parameters, such as temperature, in addition to mechanical and electromagnetic parameters. (From Hawley's Condensed Chemical Dictionary, 12th ed)	Thermodynamic
D014158	Transcription, Genetic	The biosynthesis of RNA carried out on a template of DNA. The biosynthesis of DNA from an RNA template is called REVERSE TRANSCRIPTION.	Genetic Transcription
D014443	Tyrosine	A non-essential amino acid. In animals it is synthesized from PHENYLALANINE. It is also the precursor of EPINEPHRINE; THYROID HORMONES; and melanin.	L-Tyrosine,Tyrosine, L-isomer,para-Tyrosine,L Tyrosine,Tyrosine, L isomer,para Tyrosine
D014448	Tyrosine-tRNA Ligase	An enzyme that activates tyrosine with its specific transfer RNA. EC 6.1.1.1.	Tyrosyl T RNA Synthetase,Tyr-tRNA Ligase,Tyrosyl-tRNA Synthetase,Ligase, Tyr-tRNA,Ligase, Tyrosine-tRNA,Synthetase, Tyrosyl-tRNA,Tyr tRNA Ligase,Tyrosine tRNA Ligase,Tyrosyl tRNA Synthetase
D015870	Gene Expression	The phenotypic manifestation of a gene or genes by the processes of GENETIC TRANSCRIPTION and GENETIC TRANSLATION.	Expression, Gene,Expressions, Gene,Gene Expressions