Glycyl‐tRNA synthetase from <i>Thermus thermophilus</i>

Mazauric, Marie‐Hélène; Keith, Gérard; Logan, D.T.; Kreutzer, Roland; Giegé, Richard; Kern, Daniel

doi:10.1046/j.1432-1327.1998.2510744.x

Cited by 31 publications

(49 citation statements)

References 40 publications

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“…This glycyl-tRNA synthetase is similar to the archaeal and eukaryotic enzymes, which are dimeric, but is highly divergent from other prokaryotic enzymes, such as that of E. coli, which are tetrameric (22). E. coli glycyl-tRNA synthetase can charge only prokaryotic tRNA Gly , whereas the enzyme from T. thermophilus is also able to charge eukaryotic tRNAGly .…”

Section: Resultsmentioning

confidence: 89%

The Accessory Subunit of Xenopus laevis Mitochondrial DNA Polymerase γ Increases Processivity of the Catalytic Subunit of Human DNA Polymerase γ and Is Related to Class II Aminoacyl-tRNA Synthetases

Carrodeguas

Kobayashi

Lim

et al. 1999

Molecular and Cellular Biology

102

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Peptide sequences obtained from the accessory subunit of Xenopus laevis mitochondrial DNA (mtDNA) polymerase ␥ (pol ␥) were used to clone the cDNA encoding this protein. Amino-terminal sequencing of the mitochondrial protein indicated the presence of a 44-amino-acid mitochondrial targeting sequence, leaving a predicted mature protein with 419 amino acids and a molecular mass of 47.3 kDa. This protein is associated with the larger, catalytic subunit in preparations of active mtDNA polymerase. The small subunit exhibits homology to its human, mouse, and Drosophila counterparts. Interestingly, significant homology to glycyl-tRNA synthetases from prokaryotic organisms reveals a likely evolutionary relationship. Since attempts to produce an enzymatically active recombinant catalytic subunit of Xenopus DNA pol ␥ have not been successful, we tested the effects of adding the small subunit of the Xenopus enzyme to the catalytic subunit of human DNA pol ␥ purified from baculovirus-infected insect cells. These experiments provide the first functional evidence that the small subunit of DNA pol ␥ stimulates processive DNA synthesis by the human catalytic subunit under physiological salt conditions. Mitochondrial DNA (mtDNA) is replicated by a DNA polymerase, DNA polymerase ␥ (pol ␥), that is distinct from nuclear DNA polymerases ␣, ␤, ␦, ε, and . Since DNA pol ␥ represents only a small fraction of total cellular DNA polymerase, purification and characterization of the subunit composition of this enzyme have been difficult. Molecular cloning has contributed greatly to understanding the structure of DNA pol ␥ in different organisms. The catalytic subunits of DNA pol ␥ have been cloned for several organisms (6,16,17,27,34) and have been found to resemble family A of DNA polymerases, related to Escherichia coli DNA pol I. In Saccharomyces cerevisiae DNA pol ␥ is composed of a single polypeptide, while in Drosophila melanogaster DNA pol ␥ is comprised of two different polypeptides, a catalytic subunit of 125 kDa and an accessory subunit of 41 kDa (24, 33). The Drosophila subunits copurify and have been shown to interact, but the recombinant proteins have not yet been shown to be functional. The function of the small subunit, which we refer to as pol ␥B, is unknown. It has been proposed to influence the processivity of the catalytic subunit. Putative mammalian homologs of the Drosophila accessory subunit have been identified in sequence databases. One published purification scheme for human DNA pol ␥ suggested the existence of a small subunit (11), but the potential relationship between this polypeptide and Drosophila pol ␥B has not been established. Recently, the catalytic subunit of human DNA pol ␥ was expressed in an active form (10,19). Surprisingly, the recombinant catalytic subunit alone displayed most of the characteristics of the enzyme purified from human cells, which did not appear to contain a stoichiometric amount of a small subunit. Thus, it is not clear what role, if any, is played by putative human DNA pol ␥B.We hav...

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Section: Resultsmentioning

confidence: 89%

The Accessory Subunit of Xenopus laevis Mitochondrial DNA Polymerase γ Increases Processivity of the Catalytic Subunit of Human DNA Polymerase γ and Is Related to Class II Aminoacyl-tRNA Synthetases

Carrodeguas

Kobayashi

Lim

et al. 1999

Molecular and Cellular Biology

102

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“…Gly from both bacteria and eukaryotes, whereas mammalian GlyRS can charge only eukaryotic tRNA Gly (17). Thus, insertion III could provide additional tRNA recognition that is eukaryotespecific.…”

Section: Resultsmentioning

confidence: 99%

Long-range structural effects of a Charcot–Marie–Tooth disease-causing mutation in human glycyl-tRNA synthetase

Xie

Nangle

Zhang

et al. 2007

Proc. Natl. Acad. Sci. U.S.A.

108

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Functional expansion of specific tRNA synthetases in higher organisms is well documented. These additional functions may explain why dominant mutations in glycyl-tRNA synthetase (GlyRS) and tyrosyl-tRNA synthetase cause Charcot-Marie-Tooth (CMT) disease, the most common heritable disease of the peripheral nervous system. At least 10 disease-causing mutant alleles of GlyRS have been annotated. These mutations scatter broadly across the primary sequence and have no apparent unifying connection. Here we report the structure of wild type and a CMT-causing mutant (G526R) of homodimeric human GlyRS. The mutation is at the site for synthesis of glycyl-adenylate, but the rest of the two structures are closely similar. Significantly, the mutant form diffracts to a higher resolution and has a greater dimer interface. The extra dimer interactions are located Ϸ30 Å away from the G526R mutation. Direct experiments confirm the tighter dimer interaction of the G526R protein. The results suggest the possible importance of subtle, long-range structural effects of CMT-causing mutations at the dimer interface. From analysis of a third crystal, an appended motif, found in higher eukaryote GlyRSs, seems not to have a role in these long-range effects.crystal structure ͉ structure-function analysis ͉ dimer interface ͉ inherited peripheral neuropathy ͉ aminoacyl tRNA synthetase

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“…The discriminator base is typically A in archaeal and eukaryotic tRNA Gly and usually U in bacterial examples, where in the E. coli dimeric GlyRS it is a critical identity element (50). However, the dimeric Thermus thermophilus GlyRS is able to efficiently glycylate …”

Section: Transplantation Of Sep Identity Elements Into E Coli Trna Gmentioning

confidence: 99%

Emergence of the universal genetic code imprinted in an RNA record

Hohn¹,

Park²,

O’Donoghue³

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

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The molecular basis of the genetic code manifests itself in the interaction of the aminoacyl-tRNA synthetases and their cognate tRNAs. The fundamental biological question regarding these enzymes' role in the evolution of the genetic code remains open. Here we probe this question in a system in which the same tRNA species is aminoacylated by two unrelated synthetases. Should this tRNA possess major identity elements common to both enzymes, this would favor a scenario where the aminoacyl-tRNA synthetases evolved in the context of preestablished tRNA identity, i.e., after the universal genetic code emerged. An experimental system is provided by the recently discovered O-phosphoseryl-tRNA synthetase (SepRS), which acylates tRNA Cys with phosphoserine (Sep), and the well known cysteinyl-tRNA synthetase, which charges the same tRNA with cysteine. We determined the identity elements of Methanocaldococcus jannaschii tRNA Cys in the aminoacylation reaction for the two Methanococcus maripaludis synthetases SepRS (forming Sep-tRNA Cys ) and cysteinyl-tRNA synthetase (forming Cys-tRNA Cys ). The major elements, the discriminator base and the three anticodon bases, are shared by both tRNA synthetases. An evolutionary analysis of archaeal, bacterial, and eukaryotic tRNA Cys sequences predicted additional SepRS-specific minor identity elements (G37, A47, and A59) and suggested the dominance of vertical inheritance for tRNA Cys from a single common ancestor. Transplantation of the identified identity elements into the Escherichia coli tRNA Gly scaffold endowed facile phosphoserylation activity on the resulting chimera. Thus, tRNA Cys identity is an ancient RNA record that depicts the emergence of the universal genetic code before the evolution of the modern aminoacylation systems.aminoacyl-tRNA synthetase ͉ evolution ͉ O-phosphoseryl-tRNA synthetase ͉ tRNA identity

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Glycyl‐tRNA synthetase from Thermus thermophilus

Cited by 31 publications

References 40 publications

The Accessory Subunit of Xenopus laevis Mitochondrial DNA Polymerase γ Increases Processivity of the Catalytic Subunit of Human DNA Polymerase γ and Is Related to Class II Aminoacyl-tRNA Synthetases

The Accessory Subunit of Xenopus laevis Mitochondrial DNA Polymerase γ Increases Processivity of the Catalytic Subunit of Human DNA Polymerase γ and Is Related to Class II Aminoacyl-tRNA Synthetases

Long-range structural effects of a Charcot–Marie–Tooth disease-causing mutation in human glycyl-tRNA synthetase

Emergence of the universal genetic code imprinted in an RNA record

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