The Experts below are selected from a list of 237 Experts worldwide ranked by ideXlab platform
Gregor Meyers - One of the best experts on this subject based on the ideXlab platform.
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structure function relationship in the termination upstream Ribosomal Binding Site of the calicivirus rabbit hemorrhagic disease virus
Nucleic Acids Research, 2019Co-Authors: Rene Wennesz, Christine Luttermann, Felix Kreher, Gregor MeyersAbstract:Caliciviruses use a termination/reinitiation mechanism for translation of their minor capsid protein VP2. A sequence element of about 80 nucleotides denoted 'termination upstream Ribosomal Binding Site' (TURBS) is crucial for reinitiation. RNA secondary structure probing and computer aided secondary structure prediction revealed a rather low degree of secondary structure determinants for the TURBS of the rabbit hermorrhagic disease virus. Mutation analysis showed that prevention of duplex formation had major impact on the VP2 expression levels. Restoration of complementarity of the respective sequences by reciprocal mutation at least partially restored reinitiating rates. Synthetic TURBS structures preserving only the secondary structure forming sequences and the known short motifs important for TURBS function were found to drive reinitiation when the altered sequence could be predicted to allow establishment of the crucial secondary structures of the TURBS.
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feline calicivirus can tolerate gross changes of its minor capsid protein expression levels induced by changing translation reinitiation frequency or use of a separate vp2 coding mrna
PLOS ONE, 2014Co-Authors: Christine Luttermann, Gregor MeyersAbstract:Caliciviruses use reinitiation of translation governed by a ‘termination upstream Ribosomal Binding Site’ (TURBS) for expression of their minor capsid protein VP2. Mutation analysis allowed to identify sequences surrounding the translational start/stop Site of the feline calicivirus (FCV) that fine tune reinitiation frequency. A selection of these changes was introduced into the infectious FCV cDNA clone to check the influence of altered VP2 levels on virus replication. In addition, full length constructs were established that displayed a conformation, in which VP2 expression occurred under control of a duplicated subgenomic promoter. Viable viruses recovered from such constructs revealed a rather broad range of VP2 expression levels but comparable growth kinetics showing that caliciviruses can tolerate gross changes of the VP2 expression level.
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Cis-Acting RNA Structures Promoting Translation Reinitiation in Caliciviruses
2012Co-Authors: Gregor Meyers, Christine Luttermann, Maria Haß, René WenneszAbstract:Caliciviruses use a termination/reinitiation mechanism for expression of their minor capsid protein VP2. This process is dependent on defined sequence motifs in a region located upstream of the VP2 translational start Site which is called the ‘termination upstream Ribosomal Binding Site’ (TURBS). This element is necessary to tether the posttermination ribosome to the viral RNA to allow reloading of initiation factors and restart of translation. In general, the molecular principles of this mechanism show similarity to prokaryotic translation initiation. The talk will summarize basic features of this interesting mechanism of gene expression and present the results of RNA structure and function relationship of the TURBS
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the importance of inter and intramolecular base pairing for translation reinitiation on a eukaryotic bicistronic mrna
Genes & Development, 2009Co-Authors: Christine Luttermann, Gregor MeyersAbstract:Calicivirus structure proteins are expressed from a subgenomic mRNA with two overlapping cistrons. The first ORF of this RNA codes for the viral major capsid protein VP1, and the second for the minor capsid protein VP2. Translation of VP2 is mediated by a termination/reinitiation mechanism, which depends on an upstream sequence element of ∼70 nucleotides denoted “termination upstream Ribosomal Binding Site” (TURBS). Two short sequence motifs within the TURBS were found to be essential for reinitiation. By a whole set of single Site mutations and reciprocal base exchanges we demonstrate here for the first time conclusive evidence for the necessity of mRNA/18S rRNA hybridization for translation reinitiation in an eukaryotic system. Moreover, we show that motif 2 exhibits intramolecular hybridization with a complementary region upstream of motif 1, thus forming a secondary structure that positions post-termination ribosomes in an optimal distance to the VP2 start codon. Analysis of the essential elements of the TURBS led to a better understanding of the requirements for translation termination/reinitiation in eukaryotes.
Pascale Romby - One of the best experts on this subject based on the ideXlab platform.
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the escherichia coli threonyl trna synthetase gene contains a split Ribosomal Binding Site interrupted by a hairpin structure that is essential for autoregulation
Molecular Microbiology, 1998Co-Authors: Christine Sacerdot, M. Graffe, Joël Caillet, Chantal Ehresmann, Bernard Ehresmann, Flore Eyermann, Mathias Springer, Pascale RombyAbstract:Summary The expression of the gene encoding Escherichia coli threonyl-tRNA synthetase (ThrRS) is negatively autoregulated at the translational level. ThrRS binds to its own mRNA leader, which consists of four structural and functional domains: the Shine‐Dalgarno (SD) sequence and the initiation codon region (domain 1); two upstream hairpins (domains 2 and 4) connected by a single-stranded region (domain 3). Using a combination of in vivo and in vitro approaches, we show here that the ribosome binds to thrS mRNA at two non-contiguous Sites: region π12 to ˛16 comprising the SD sequence and the AUG codon and, unexpectedly, an upstream single-stranded sequence in domain 3. These two regions are brought into close proximity by a 38-nucleotide-long hairpin structure (domain 2). This domain, although adjacent to the 58 edge of the SD sequence, does not inhibit ribosome Binding as long as the single-stranded region of domain 3 is present. A stretch of unpaired nucleotides in domain 3, but not a specific sequence, is required for efficient translation. As the repressor and the ribosome bind to interspersed domains, the competition between ThrRS and ribosome for thrS mRNA Binding can be explained by steric hindrance.
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The Escherichia coli threonyl‐tRNA synthetase gene contains a split Ribosomal Binding Site interrupted by a hairpin structure that is essential for autoregulation
Molecular microbiology, 1998Co-Authors: Christine Sacerdot, M. Graffe, Joël Caillet, Chantal Ehresmann, Bernard Ehresmann, Flore Eyermann, Mathias Springer, Pascale RombyAbstract:Summary The expression of the gene encoding Escherichia coli threonyl-tRNA synthetase (ThrRS) is negatively autoregulated at the translational level. ThrRS binds to its own mRNA leader, which consists of four structural and functional domains: the Shine‐Dalgarno (SD) sequence and the initiation codon region (domain 1); two upstream hairpins (domains 2 and 4) connected by a single-stranded region (domain 3). Using a combination of in vivo and in vitro approaches, we show here that the ribosome binds to thrS mRNA at two non-contiguous Sites: region π12 to ˛16 comprising the SD sequence and the AUG codon and, unexpectedly, an upstream single-stranded sequence in domain 3. These two regions are brought into close proximity by a 38-nucleotide-long hairpin structure (domain 2). This domain, although adjacent to the 58 edge of the SD sequence, does not inhibit ribosome Binding as long as the single-stranded region of domain 3 is present. A stretch of unpaired nucleotides in domain 3, but not a specific sequence, is required for efficient translation. As the repressor and the ribosome bind to interspersed domains, the competition between ThrRS and ribosome for thrS mRNA Binding can be explained by steric hindrance.
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Domains of the Escherichia coli threonyl-tRNA synthetase translational operator and their relation to threonine tRNA isoacceptors.
Journal of molecular biology, 1992Co-Authors: Claude Brunel, M. Graffe, J. Dondon, Joël Caillet, Pascale Lesage, H. Moine, Pascale Romby, Chantal Ehresmann, Bernard Ehresmann, Marianne Grunberg-managoAbstract:Abstract The expression of the gene for threonyl-tRNA synthetase ( thrS ) is negatively autoregulated at the translational level in Escherichia coli . The synthetase binds to a region of the thrS leader mRNA upstream from the Ribosomal Binding Site inhibiting subsequent translation. The leader mRNA consists of four structural domains. The present work shows that mutations in these four domains affect expression and/or regulation in different ways. Domain 1, the 3′ end of the leader, contains the Ribosomal Binding Site, which appears not to be essential for synthetase Binding. Mutations in this domain probably affect regulation by changing the competition between the ribosome and the synthetase for Binding to the leader. Domain 2, 3′ from the Ribosomal Binding Site, is a stem and loop with structural similarities to the tRNA Thr anticodon arm. In tRNAs the anticodon loop is seven nucleotides long, mutations that increase or decrease the length of the anticodon-like loop of domain 2 from seven nucleotides abolish control. The nucleotides in the second and third positions of the anticodon-like sequence are essential for recognition and the nucleotide in the wobble position is not, again like tRNA Thr . The effect of mutations in domain 3 indicate that it acts as an articulation between domains 2 and 4. Domain 4 is a stable arm that has similarities to the acceptor arm of tRNA Thr and is shown to be necessary for regulation. Based on this mutational analysis and previous footprinting experiments, it appears that domains 2 and 4, those analogous to tRNA Thr , are involved in Binding the synthetase which inhibits translation probably by interfering with ribosome loading at the nearby translation initiation Site.
M. Graffe - One of the best experts on this subject based on the ideXlab platform.
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the escherichia coli threonyl trna synthetase gene contains a split Ribosomal Binding Site interrupted by a hairpin structure that is essential for autoregulation
Molecular Microbiology, 1998Co-Authors: Christine Sacerdot, M. Graffe, Joël Caillet, Chantal Ehresmann, Bernard Ehresmann, Flore Eyermann, Mathias Springer, Pascale RombyAbstract:Summary The expression of the gene encoding Escherichia coli threonyl-tRNA synthetase (ThrRS) is negatively autoregulated at the translational level. ThrRS binds to its own mRNA leader, which consists of four structural and functional domains: the Shine‐Dalgarno (SD) sequence and the initiation codon region (domain 1); two upstream hairpins (domains 2 and 4) connected by a single-stranded region (domain 3). Using a combination of in vivo and in vitro approaches, we show here that the ribosome binds to thrS mRNA at two non-contiguous Sites: region π12 to ˛16 comprising the SD sequence and the AUG codon and, unexpectedly, an upstream single-stranded sequence in domain 3. These two regions are brought into close proximity by a 38-nucleotide-long hairpin structure (domain 2). This domain, although adjacent to the 58 edge of the SD sequence, does not inhibit ribosome Binding as long as the single-stranded region of domain 3 is present. A stretch of unpaired nucleotides in domain 3, but not a specific sequence, is required for efficient translation. As the repressor and the ribosome bind to interspersed domains, the competition between ThrRS and ribosome for thrS mRNA Binding can be explained by steric hindrance.
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The Escherichia coli threonyl‐tRNA synthetase gene contains a split Ribosomal Binding Site interrupted by a hairpin structure that is essential for autoregulation
Molecular microbiology, 1998Co-Authors: Christine Sacerdot, M. Graffe, Joël Caillet, Chantal Ehresmann, Bernard Ehresmann, Flore Eyermann, Mathias Springer, Pascale RombyAbstract:Summary The expression of the gene encoding Escherichia coli threonyl-tRNA synthetase (ThrRS) is negatively autoregulated at the translational level. ThrRS binds to its own mRNA leader, which consists of four structural and functional domains: the Shine‐Dalgarno (SD) sequence and the initiation codon region (domain 1); two upstream hairpins (domains 2 and 4) connected by a single-stranded region (domain 3). Using a combination of in vivo and in vitro approaches, we show here that the ribosome binds to thrS mRNA at two non-contiguous Sites: region π12 to ˛16 comprising the SD sequence and the AUG codon and, unexpectedly, an upstream single-stranded sequence in domain 3. These two regions are brought into close proximity by a 38-nucleotide-long hairpin structure (domain 2). This domain, although adjacent to the 58 edge of the SD sequence, does not inhibit ribosome Binding as long as the single-stranded region of domain 3 is present. A stretch of unpaired nucleotides in domain 3, but not a specific sequence, is required for efficient translation. As the repressor and the ribosome bind to interspersed domains, the competition between ThrRS and ribosome for thrS mRNA Binding can be explained by steric hindrance.
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Stabilised Secondary Structure at a Ribosomal Binding Site Enhances Translational Repression inE. Coli
Journal of molecular biology, 1995Co-Authors: C. Brunel, P. Romby, C. Sacerdot, M H De Smit, M. Graffe, J. Dondon, J. Van Duin, B. Ehresmann, C. Ehresmann, M. SpringerAbstract:Abstract The expression of the gene encoding Escherichia coli threonyl-tRNA synthetase is negatively autoregulated at the translational level. The negative feedback is due to the Binding of the synthetase to an operator Site on its own mRNA located upstream of the initiation codon. The present work describes the characterisation of operator mutants that have the rare property of enhancing repression. These mutations cause (1) a low basal level of expression, (2) a temperature-dependent expression, and (3) an increased capacity of the synthetase to repress its own expression at low temperature. Surprisingly, this enhancement of repression is not explained by an increase of affinity of the mutant operators for the enzyme but by the formation, at low temperature, of a few supplementary base-pairs between the Ribosomal Binding Site and a normally single-stranded domain of the operator. Although this additional base-pairing only slightly inhibits ribosome Binding in the absence of repressor, simple thermodynamic considerations indicate that this is sufficient to increase repression. This increase is explained by the competition between the ribosome and repressor for overlapping regions of the mRNA. When the Ribosomal Binding Site is base-paired, the ribosome cannot bind while the repressor can, giving the repressor the advantage in the competition. Thus, the existence of an open versus base-paired equilibrium in a Ribosomal Binding Site of a translational operator amplifies the magnitude of control. This molecular amplification device might be an essential component of translational control considering the low free repressor/ribosome ratio of the low affinity of translational repressors for their target operators.
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Domains of the Escherichia coli threonyl-tRNA synthetase translational operator and their relation to threonine tRNA isoacceptors.
Journal of molecular biology, 1992Co-Authors: Claude Brunel, M. Graffe, J. Dondon, Joël Caillet, Pascale Lesage, H. Moine, Pascale Romby, Chantal Ehresmann, Bernard Ehresmann, Marianne Grunberg-managoAbstract:Abstract The expression of the gene for threonyl-tRNA synthetase ( thrS ) is negatively autoregulated at the translational level in Escherichia coli . The synthetase binds to a region of the thrS leader mRNA upstream from the Ribosomal Binding Site inhibiting subsequent translation. The leader mRNA consists of four structural domains. The present work shows that mutations in these four domains affect expression and/or regulation in different ways. Domain 1, the 3′ end of the leader, contains the Ribosomal Binding Site, which appears not to be essential for synthetase Binding. Mutations in this domain probably affect regulation by changing the competition between the ribosome and the synthetase for Binding to the leader. Domain 2, 3′ from the Ribosomal Binding Site, is a stem and loop with structural similarities to the tRNA Thr anticodon arm. In tRNAs the anticodon loop is seven nucleotides long, mutations that increase or decrease the length of the anticodon-like loop of domain 2 from seven nucleotides abolish control. The nucleotides in the second and third positions of the anticodon-like sequence are essential for recognition and the nucleotide in the wobble position is not, again like tRNA Thr . The effect of mutations in domain 3 indicate that it acts as an articulation between domains 2 and 4. Domain 4 is a stable arm that has similarities to the acceptor arm of tRNA Thr and is shown to be necessary for regulation. Based on this mutational analysis and previous footprinting experiments, it appears that domains 2 and 4, those analogous to tRNA Thr , are involved in Binding the synthetase which inhibits translation probably by interfering with ribosome loading at the nearby translation initiation Site.
Christine Luttermann - One of the best experts on this subject based on the ideXlab platform.
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structure function relationship in the termination upstream Ribosomal Binding Site of the calicivirus rabbit hemorrhagic disease virus
Nucleic Acids Research, 2019Co-Authors: Rene Wennesz, Christine Luttermann, Felix Kreher, Gregor MeyersAbstract:Caliciviruses use a termination/reinitiation mechanism for translation of their minor capsid protein VP2. A sequence element of about 80 nucleotides denoted 'termination upstream Ribosomal Binding Site' (TURBS) is crucial for reinitiation. RNA secondary structure probing and computer aided secondary structure prediction revealed a rather low degree of secondary structure determinants for the TURBS of the rabbit hermorrhagic disease virus. Mutation analysis showed that prevention of duplex formation had major impact on the VP2 expression levels. Restoration of complementarity of the respective sequences by reciprocal mutation at least partially restored reinitiating rates. Synthetic TURBS structures preserving only the secondary structure forming sequences and the known short motifs important for TURBS function were found to drive reinitiation when the altered sequence could be predicted to allow establishment of the crucial secondary structures of the TURBS.
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feline calicivirus can tolerate gross changes of its minor capsid protein expression levels induced by changing translation reinitiation frequency or use of a separate vp2 coding mrna
PLOS ONE, 2014Co-Authors: Christine Luttermann, Gregor MeyersAbstract:Caliciviruses use reinitiation of translation governed by a ‘termination upstream Ribosomal Binding Site’ (TURBS) for expression of their minor capsid protein VP2. Mutation analysis allowed to identify sequences surrounding the translational start/stop Site of the feline calicivirus (FCV) that fine tune reinitiation frequency. A selection of these changes was introduced into the infectious FCV cDNA clone to check the influence of altered VP2 levels on virus replication. In addition, full length constructs were established that displayed a conformation, in which VP2 expression occurred under control of a duplicated subgenomic promoter. Viable viruses recovered from such constructs revealed a rather broad range of VP2 expression levels but comparable growth kinetics showing that caliciviruses can tolerate gross changes of the VP2 expression level.
-
Cis-Acting RNA Structures Promoting Translation Reinitiation in Caliciviruses
2012Co-Authors: Gregor Meyers, Christine Luttermann, Maria Haß, René WenneszAbstract:Caliciviruses use a termination/reinitiation mechanism for expression of their minor capsid protein VP2. This process is dependent on defined sequence motifs in a region located upstream of the VP2 translational start Site which is called the ‘termination upstream Ribosomal Binding Site’ (TURBS). This element is necessary to tether the posttermination ribosome to the viral RNA to allow reloading of initiation factors and restart of translation. In general, the molecular principles of this mechanism show similarity to prokaryotic translation initiation. The talk will summarize basic features of this interesting mechanism of gene expression and present the results of RNA structure and function relationship of the TURBS
-
the importance of inter and intramolecular base pairing for translation reinitiation on a eukaryotic bicistronic mrna
Genes & Development, 2009Co-Authors: Christine Luttermann, Gregor MeyersAbstract:Calicivirus structure proteins are expressed from a subgenomic mRNA with two overlapping cistrons. The first ORF of this RNA codes for the viral major capsid protein VP1, and the second for the minor capsid protein VP2. Translation of VP2 is mediated by a termination/reinitiation mechanism, which depends on an upstream sequence element of ∼70 nucleotides denoted “termination upstream Ribosomal Binding Site” (TURBS). Two short sequence motifs within the TURBS were found to be essential for reinitiation. By a whole set of single Site mutations and reciprocal base exchanges we demonstrate here for the first time conclusive evidence for the necessity of mRNA/18S rRNA hybridization for translation reinitiation in an eukaryotic system. Moreover, we show that motif 2 exhibits intramolecular hybridization with a complementary region upstream of motif 1, thus forming a secondary structure that positions post-termination ribosomes in an optimal distance to the VP2 start codon. Analysis of the essential elements of the TURBS led to a better understanding of the requirements for translation termination/reinitiation in eukaryotes.
Bernard Ehresmann - One of the best experts on this subject based on the ideXlab platform.
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the escherichia coli threonyl trna synthetase gene contains a split Ribosomal Binding Site interrupted by a hairpin structure that is essential for autoregulation
Molecular Microbiology, 1998Co-Authors: Christine Sacerdot, M. Graffe, Joël Caillet, Chantal Ehresmann, Bernard Ehresmann, Flore Eyermann, Mathias Springer, Pascale RombyAbstract:Summary The expression of the gene encoding Escherichia coli threonyl-tRNA synthetase (ThrRS) is negatively autoregulated at the translational level. ThrRS binds to its own mRNA leader, which consists of four structural and functional domains: the Shine‐Dalgarno (SD) sequence and the initiation codon region (domain 1); two upstream hairpins (domains 2 and 4) connected by a single-stranded region (domain 3). Using a combination of in vivo and in vitro approaches, we show here that the ribosome binds to thrS mRNA at two non-contiguous Sites: region π12 to ˛16 comprising the SD sequence and the AUG codon and, unexpectedly, an upstream single-stranded sequence in domain 3. These two regions are brought into close proximity by a 38-nucleotide-long hairpin structure (domain 2). This domain, although adjacent to the 58 edge of the SD sequence, does not inhibit ribosome Binding as long as the single-stranded region of domain 3 is present. A stretch of unpaired nucleotides in domain 3, but not a specific sequence, is required for efficient translation. As the repressor and the ribosome bind to interspersed domains, the competition between ThrRS and ribosome for thrS mRNA Binding can be explained by steric hindrance.
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The Escherichia coli threonyl‐tRNA synthetase gene contains a split Ribosomal Binding Site interrupted by a hairpin structure that is essential for autoregulation
Molecular microbiology, 1998Co-Authors: Christine Sacerdot, M. Graffe, Joël Caillet, Chantal Ehresmann, Bernard Ehresmann, Flore Eyermann, Mathias Springer, Pascale RombyAbstract:Summary The expression of the gene encoding Escherichia coli threonyl-tRNA synthetase (ThrRS) is negatively autoregulated at the translational level. ThrRS binds to its own mRNA leader, which consists of four structural and functional domains: the Shine‐Dalgarno (SD) sequence and the initiation codon region (domain 1); two upstream hairpins (domains 2 and 4) connected by a single-stranded region (domain 3). Using a combination of in vivo and in vitro approaches, we show here that the ribosome binds to thrS mRNA at two non-contiguous Sites: region π12 to ˛16 comprising the SD sequence and the AUG codon and, unexpectedly, an upstream single-stranded sequence in domain 3. These two regions are brought into close proximity by a 38-nucleotide-long hairpin structure (domain 2). This domain, although adjacent to the 58 edge of the SD sequence, does not inhibit ribosome Binding as long as the single-stranded region of domain 3 is present. A stretch of unpaired nucleotides in domain 3, but not a specific sequence, is required for efficient translation. As the repressor and the ribosome bind to interspersed domains, the competition between ThrRS and ribosome for thrS mRNA Binding can be explained by steric hindrance.
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Domains of the Escherichia coli threonyl-tRNA synthetase translational operator and their relation to threonine tRNA isoacceptors.
Journal of molecular biology, 1992Co-Authors: Claude Brunel, M. Graffe, J. Dondon, Joël Caillet, Pascale Lesage, H. Moine, Pascale Romby, Chantal Ehresmann, Bernard Ehresmann, Marianne Grunberg-managoAbstract:Abstract The expression of the gene for threonyl-tRNA synthetase ( thrS ) is negatively autoregulated at the translational level in Escherichia coli . The synthetase binds to a region of the thrS leader mRNA upstream from the Ribosomal Binding Site inhibiting subsequent translation. The leader mRNA consists of four structural domains. The present work shows that mutations in these four domains affect expression and/or regulation in different ways. Domain 1, the 3′ end of the leader, contains the Ribosomal Binding Site, which appears not to be essential for synthetase Binding. Mutations in this domain probably affect regulation by changing the competition between the ribosome and the synthetase for Binding to the leader. Domain 2, 3′ from the Ribosomal Binding Site, is a stem and loop with structural similarities to the tRNA Thr anticodon arm. In tRNAs the anticodon loop is seven nucleotides long, mutations that increase or decrease the length of the anticodon-like loop of domain 2 from seven nucleotides abolish control. The nucleotides in the second and third positions of the anticodon-like sequence are essential for recognition and the nucleotide in the wobble position is not, again like tRNA Thr . The effect of mutations in domain 3 indicate that it acts as an articulation between domains 2 and 4. Domain 4 is a stable arm that has similarities to the acceptor arm of tRNA Thr and is shown to be necessary for regulation. Based on this mutational analysis and previous footprinting experiments, it appears that domains 2 and 4, those analogous to tRNA Thr , are involved in Binding the synthetase which inhibits translation probably by interfering with ribosome loading at the nearby translation initiation Site.
