Identification of Amino Acids in the N-terminal Domain of Atypical Methanogenic-type Seryl-tRNA Synthetase Critical for tRNA Recognition
2009; Elsevier BV; Volume: 284; Issue: 44 Linguagem: Inglês
10.1074/jbc.m109.044099
ISSN1083-351X
AutoresJelena Jarić, Silvija Bilokapić, Sonja Lesjak, Ana Crnković, Nenad Ban, Ivana Weygand-Đurašević,
Tópico(s)Genomics and Phylogenetic Studies
ResumoSeryl-tRNA synthetase (SerRS) from methanogenic archaeon Methanosarcina barkeri, contains an idiosyncratic N-terminal domain, composed of an antiparallel β-sheet capped by a helical bundle, connected to the catalytic core by a short linker peptide. It is very different from the coiled-coil tRNA binding domain in bacterial-type SerRS. Because the crystal structure of the methanogenic-type SerRS·tRNA complex has not been obtained, a docking model was produced, which indicated that highly conserved helices H2 and H3 of the N-terminal domain may be important for recognition of the extra arm of tRNASer. Based on structural information and the docking model, we have mutated various positions within the N-terminal region and probed their involvement in tRNA binding and serylation. Total loss of activity and inability of the R76A variant to form the complex with cognate tRNA identifies Arg76 located in helix H2 as a crucial tRNA-interacting residue. Alteration of Lys79 positioned in helix H2 and Arg94 in the loop between helix H2 and β-strand A4 have a pronounced effect on SerRS·tRNASer complex formation and dissociation constants (KD) determined by surface plasmon resonance. The replacement of residues Arg38 (located in the loop between helix H1 and β-strand A2), Lys141 and Asn142 (from H3), and Arg143 (between H3 and H4) moderately affect both the serylation activity and the KD values. Furthermore, we have obtained a striking correlation between these results and in vivo effects of these mutations by quantifying the efficiency of suppression of bacterial amber mutations, after coexpression of the genes for M. barkeri suppressor tRNASer and a set of mMbSerRS variants in Escherichia coli. Seryl-tRNA synthetase (SerRS) from methanogenic archaeon Methanosarcina barkeri, contains an idiosyncratic N-terminal domain, composed of an antiparallel β-sheet capped by a helical bundle, connected to the catalytic core by a short linker peptide. It is very different from the coiled-coil tRNA binding domain in bacterial-type SerRS. Because the crystal structure of the methanogenic-type SerRS·tRNA complex has not been obtained, a docking model was produced, which indicated that highly conserved helices H2 and H3 of the N-terminal domain may be important for recognition of the extra arm of tRNASer. Based on structural information and the docking model, we have mutated various positions within the N-terminal region and probed their involvement in tRNA binding and serylation. Total loss of activity and inability of the R76A variant to form the complex with cognate tRNA identifies Arg76 located in helix H2 as a crucial tRNA-interacting residue. Alteration of Lys79 positioned in helix H2 and Arg94 in the loop between helix H2 and β-strand A4 have a pronounced effect on SerRS·tRNASer complex formation and dissociation constants (KD) determined by surface plasmon resonance. The replacement of residues Arg38 (located in the loop between helix H1 and β-strand A2), Lys141 and Asn142 (from H3), and Arg143 (between H3 and H4) moderately affect both the serylation activity and the KD values. Furthermore, we have obtained a striking correlation between these results and in vivo effects of these mutations by quantifying the efficiency of suppression of bacterial amber mutations, after coexpression of the genes for M. barkeri suppressor tRNASer and a set of mMbSerRS variants in Escherichia coli. The aminoacyl-tRNA synthetases (aaRSs) 2The abbreviations used are: aaRSaminoacyl-tRNA synthetase (standard amino acid abbreviations precede RS throughout)SerRSseryl-tRNA synthetasemMbSerRSmethanogenic-type Methanosarcina barkeri SerRSMbtRNAGGASerM. barkeri tRNASer with anticodon GGASPRsurface plasmon resonanceWTwild typeDTTdithiothreitolMes2-(N-morpholino)ethanesulfonic acid. 2The abbreviations used are: aaRSaminoacyl-tRNA synthetase (standard amino acid abbreviations precede RS throughout)SerRSseryl-tRNA synthetasemMbSerRSmethanogenic-type Methanosarcina barkeri SerRSMbtRNAGGASerM. barkeri tRNASer with anticodon GGASPRsurface plasmon resonanceWTwild typeDTTdithiothreitolMes2-(N-morpholino)ethanesulfonic acid. catalyze the activation of cognate amino acids and their transfer to the 3′-end of corresponding tRNA molecules. The aaRSs are a highly conserved family of enzymes comprised of two distinct structural groups referred to as classes I and II (1Ibba M. Soll D. Annu. Rev. Biochem. 2000; 69: 617-650Crossref PubMed Scopus (1076) Google Scholar, 2Eriani G. Delarue M. Poch O. Gangloff J. Moras D. Nature. 1990; 347: 203-206Crossref PubMed Scopus (1182) Google Scholar, 3Cusack S. Berthet-Colominas C. Härtlein M. Nassar N. Leberman R. Nature. 1990; 347: 249-255Crossref PubMed Scopus (563) Google Scholar), with a notable exception of LysRS representatives, which belong to both classes (4Ibba M. Morgan S. Curnow A.W. Pridmore D.R. Vothknecht U.C. Gardner W. Lin W. Woese C.R. Söll D. Science. 1997; 278: 1119-1122Crossref PubMed Scopus (174) Google Scholar). Although the catalytic mechanisms of various aaRSs are broadly similar (5Zhang C.M. Perona J.J. Ryu K. Francklyn C. Hou Y.M. J. Mol. Biol. 2006; 361: 300-311Crossref PubMed Scopus (85) Google Scholar), each enzyme has developed a high specificity in recognizing its cognate amino acid and tRNA, which is pivotal for accurate translation of the genetic code (1Ibba M. Soll D. Annu. Rev. Biochem. 2000; 69: 617-650Crossref PubMed Scopus (1076) Google Scholar). The discrimination of the amino acids is based on recognizing the differences in the size and charge of the molecules (6Ataide S.F. Ibba M. ACS Chem. Biol. 2006; 1: 285-297Crossref PubMed Scopus (38) Google Scholar). The specificity of tRNA selection depends on a set of identity determinants that are mostly located at two distal extremities: the anticodon loop and the amino acid accepting stem. In a few instances, identity elements are also found in the D-arm, T-arm, and variable loop. They can either act as positive determinants that enhance aminoacylation or negative ones that prevent aminoacylation. The recognition of tRNAs by synthetases can also be affected by the modification of particular nucleotides (7Giegé R. Puglisi J.D. Florentz C. Prog. Nucleic Acid Res. Mol. Biol. 1993; 45: 129-206Crossref PubMed Scopus (218) Google Scholar, 8Giegé R. Sissler M. Florentz C. Nucleic Acids Res. 1998; 26: 5017-5035Crossref PubMed Scopus (628) Google Scholar). AaRSs show divergent strategies for tRNA recognition. Most notably, class I and class II aaRSs (including pyrrolysyl-tRNA synthetase, see Ref. 9Nozawa K. O'Donoghue P. Gundllapalli S. Araiso Y. Ishitani R. Umehara T. Söll D. Nureki O. Nature. 2009; 457: 1163-1167Crossref PubMed Scopus (126) Google Scholar) approach tRNAs from the minor and major groove sides of the acceptor stem, respectively (10Ribas de Pouplana L. Schimmel P. Cell. 2001; 104: 191-193Abstract Full Text Full Text PDF PubMed Scopus (142) Google Scholar). Although the majority of determinants are in direct contact with cognate synthetases (8Giegé R. Sissler M. Florentz C. Nucleic Acids Res. 1998; 26: 5017-5035Crossref PubMed Scopus (628) Google Scholar), the aminoacylation fidelity is controlled by kinetic differences more than by binding affinities (11Ebel J.P. Giegé R. Bonnet J. Kern D. Befort N. Bollack C. Fasiolo F. Gangloff J. Dirheimer G. Biochimie. 1973; 55: 547-557Crossref PubMed Scopus (154) Google Scholar).Seryl-tRNA synthetases (SerRSs), which catalyze the aminoacylation of several tRNASer isoacceptors and tRNASec with serine, can be divided into two structurally different groups: bacterial-type SerRSs function in a variety of archaeal, bacterial, and eukaryotic organisms, whereas the methanogenic-type was found only in methanogenic archaea (12Kim H.S. Vothknecht U.C. Hedderich R. Celic I. Söll D. J. Bacteriol. 1998; 180: 6446-6449Crossref PubMed Google Scholar, 13Tumbula D. Vothknecht U.C. Kim H.S. Ibba M. Min B. Li T. Pelaschier J. Stathopoulos C. Becker H. Söll D. Genetics. 1999; 152: 1269-1276PubMed Google Scholar). Furthermore, based on sequence comparison (14Lenhard B. Orellana O. Ibba M. Weygand-Duraseviæ I. Nucleic Acids Res. 1999; 27: 721-729Crossref PubMed Scopus (61) Google Scholar, 15Woese C.R. Olsen G.J. Ibba M. Söll D. Microbiol. Mol. Biol. Rev. 2000; 64: 202-236Crossref PubMed Scopus (530) Google Scholar) and x-ray analyses, two subgroups of bacterial-type SerRSs were identified: one consists of the enzymes from bacterial sources, best represented by those from Thermus thermophilus (16Fujinaga M. Berthet-Colominas C. Yaremchuk A.D. Tukalo M.A. Cusack S. J. Mol. Biol. 1993; 234: 222-233Crossref PubMed Scopus (92) Google Scholar) and Escherichia coli (3Cusack S. Berthet-Colominas C. Härtlein M. Nassar N. Leberman R. Nature. 1990; 347: 249-255Crossref PubMed Scopus (563) Google Scholar), and an archaeal/eukaryal-type, structurally related to SerRS from archaeon Pyrococcus horikoshii (17Itoh Y. Sekine S. Kuroishi C. Terada T. Shirouzu M. Kuramitsu S. Yokoyama S. RNA Biol. 2008; 5: 169-177Crossref PubMed Scopus (23) Google Scholar).All SerRSs are functional homodimers with a C-terminal active site domain typical for class II aaRSs and an N-terminal domain that is responsible for binding of the long variable arm of tRNASer isoacceptors (reviewed in Ref. 18Weygand-Durasevic I. Cusack S. The Aminoacyl-tRNA Synthetases.in: Ibba M. Francklyn C. Cusack S. Landes Bioscience, Georgetown, TX2005: 177-192Google Scholar), with exception of the mammalian mitochondrial enzyme (19Chimnaronk S. Gravers Jeppesen M. Suzuki T. Nyborg J. Watanabe K. EMBO J. 2005; 24: 3369-3379Crossref PubMed Scopus (70) Google Scholar). The long variable arm of tRNASer categorizes it as one of the type 2 tRNAs, including the tRNASer and tRNALeu species (from all organisms or domains of life) and bacterial tRNATyr species (20Sprinzl M. Vassilenko K.S. Nucleic Acids Res. 2005; 33: D139-D140Crossref PubMed Scopus (349) Google Scholar). In bacterial-type SerRS, the N-terminal domain forms an antiparallel α-helical coiled-coil structure (3Cusack S. Berthet-Colominas C. Härtlein M. Nassar N. Leberman R. Nature. 1990; 347: 249-255Crossref PubMed Scopus (563) Google Scholar, 16Fujinaga M. Berthet-Colominas C. Yaremchuk A.D. Tukalo M.A. Cusack S. J. Mol. Biol. 1993; 234: 222-233Crossref PubMed Scopus (92) Google Scholar), whereas in the methanogenic-type counterpart it is significantly larger and composed of a six-stranded antiparallel β-sheet capped by a bundle of three helices (H1, H2, and H4) with up-down topology and an additional short helix (H3) that runs almost perpendicular to helix H4 (21Bilokapic S. Maier T. Ahel D. Gruic-Sovulj I. Söll D. Weygand- Durasevic I. Ban N. EMBO J. 2006; 25: 2498-2509Crossref PubMed Scopus (72) Google Scholar) (see Fig. 1). Despite pronounced structural differences between the tRNA-binding domains in two SerRS types, in each case the N-terminal domain of one subunit interacts with the extra arm of tRNASer, to position the 3′-end of tRNA into the C-terminal active site of another subunit (23Biou V. Yaremchuk A. Tukalo M. Cusack S. Science. 1994; 263: 1404-1410Crossref PubMed Scopus (422) Google Scholar, 24Vincent C. Borel F. Willison J.C. Leberman R. Härtlein M. Nucleic Acids Res. 1995; 23: 1113-1118Crossref PubMed Scopus (36) Google Scholar, 25Bilokapic S. Ivic N. Godinic-Mikulcic V. Piantanida I. Ban N. Weygand-Durasevic I. J. Biol. Chem. 2009; 284: 10706-10713Abstract Full Text Full Text PDF PubMed Scopus (12) Google Scholar). The recent crystal structure of the first archaeal/eukaryal SerRS from the archaeon P. horikoshii, and the structure-based model of the enzyme bound with the T. thermophilus and P. horikoshii tRNAsSer, suggested that the helical N-terminal domain of P. horikoshii SerRS is also involved in the binding of the extra arm of tRNA (17Itoh Y. Sekine S. Kuroishi C. Terada T. Shirouzu M. Kuramitsu S. Yokoyama S. RNA Biol. 2008; 5: 169-177Crossref PubMed Scopus (23) Google Scholar).The recognition of tRNA by SerRS relies, besides on the long extra arm, on the identity elements in tRNASer acceptor arms, achieved by the motif 2 residues. Unlike the majority of aaRSs systems, the anticodon triplet is not recognized by SerRS. The first four base pairs in the tRNA acceptor arm (G1:C72, G2:C71, A/U3:U/A70, and R4:Y69) are identity elements for bacterial SerRS, and among them, the second G2:C71 base pair is the most significant (reviewed in Ref. 18Weygand-Durasevic I. Cusack S. The Aminoacyl-tRNA Synthetases.in: Ibba M. Francklyn C. Cusack S. Landes Bioscience, Georgetown, TX2005: 177-192Google Scholar). Consistently, the crystal structure of the T. thermophilus SerRS·tRNASer complex revealed that SerRS interacts with the tRNASer acceptor stem, and Ser261 is responsible for the base-specific interaction with G2 (23Biou V. Yaremchuk A. Tukalo M. Cusack S. Science. 1994; 263: 1404-1410Crossref PubMed Scopus (422) Google Scholar, 26Cusack S. Yaremchuk A. Tukalo M. EMBO J. 1996; 15: 2834-2842Crossref PubMed Scopus (155) Google Scholar). Although the acceptor stem sequences are not well conserved among the eukaryal tRNASer isoacceptors, the discriminator base G73 is an essential identity requirement for human tRNASer and serves as an anti-determinant in lower eukaryotes (reviewed in Ref. 18Weygand-Durasevic I. Cusack S. The Aminoacyl-tRNA Synthetases.in: Ibba M. Francklyn C. Cusack S. Landes Bioscience, Georgetown, TX2005: 177-192Google Scholar). Unlike eukaryal tRNAsSer, archaeal tRNAsSer conserve the G1:C72, C2:G71, C3:G70, and G4:C69 base pairs in the acceptor stem. However, the tRNA specificity of the archaeal/eukaryal SerRS from P. horikoshii seems to depend mainly on the extra arm, but not on the acceptor stem. Indeed, this enzyme exhibits quite relaxed specificity for tRNASer recognition (17Itoh Y. Sekine S. Kuroishi C. Terada T. Shirouzu M. Kuramitsu S. Yokoyama S. RNA Biol. 2008; 5: 169-177Crossref PubMed Scopus (23) Google Scholar).Archaeon Methanosarcina barkeri, which possesses two dissimilar SerRSs, one of a bacterial- and the other of methanogenic-type, provides an excellent system for studying the evolution of tRNASer determinants. Two enzymes recognize the same set of tRNA isoacceptors in vitro (27Korencic D. Polycarpo C. Weygand-Durasevic I. Söll D. J. Biol. Chem. 2004; 279: 48780-48786Abstract Full Text Full Text PDF PubMed Scopus (35) Google Scholar). We have undertaken several approaches to elucidate the basis for their different serylation mechanisms. Kinetic analysis of variant tRNASer transcripts by the two archaeal SerRS enzymes (27Korencic D. Polycarpo C. Weygand-Durasevic I. Söll D. J. Biol. Chem. 2004; 279: 48780-48786Abstract Full Text Full Text PDF PubMed Scopus (35) Google Scholar) revealed that the length of the variable arm is a critical recognition element for both enzymes, as is the identity of the discriminator base (G73) and base pair G30:C40 in the anticodon stem. However, additional determinants were identified as being required for specific serylation by the unusual methanogenic-type enzyme, which relies on G1:C72 identity and on the number of unpaired nucleotides at the base of the variable loop. The tRNA recognition pattern by two M. barkeri SerRS may differ in vivo, because only bacterial-type SerRS complements the function of thermolabile E. coli SerRS (28Lesjak S. Weygand-Durasevic I. FEMS Microbiol. Lett. 2009; 294: 111-118Crossref PubMed Scopus (7) Google Scholar). The aminoacyl-tRNA synthetases (aaRSs) 2The abbreviations used are: aaRSaminoacyl-tRNA synthetase (standard amino acid abbreviations precede RS throughout)SerRSseryl-tRNA synthetasemMbSerRSmethanogenic-type Methanosarcina barkeri SerRSMbtRNAGGASerM. barkeri tRNASer with anticodon GGASPRsurface plasmon resonanceWTwild typeDTTdithiothreitolMes2-(N-morpholino)ethanesulfonic acid. 2The abbreviations used are: aaRSaminoacyl-tRNA synthetase (standard amino acid abbreviations precede RS throughout)SerRSseryl-tRNA synthetasemMbSerRSmethanogenic-type Methanosarcina barkeri SerRSMbtRNAGGASerM. barkeri tRNASer with anticodon GGASPRsurface plasmon resonanceWTwild typeDTTdithiothreitolMes2-(N-morpholino)ethanesulfonic acid. catalyze the activation of cognate amino acids and their transfer to the 3′-end of corresponding tRNA molecules. The aaRSs are a highly conserved family of enzymes comprised of two distinct structural groups referred to as classes I and II (1Ibba M. Soll D. Annu. Rev. Biochem. 2000; 69: 617-650Crossref PubMed Scopus (1076) Google Scholar, 2Eriani G. Delarue M. Poch O. Gangloff J. Moras D. Nature. 1990; 347: 203-206Crossref PubMed Scopus (1182) Google Scholar, 3Cusack S. Berthet-Colominas C. Härtlein M. Nassar N. Leberman R. Nature. 1990; 347: 249-255Crossref PubMed Scopus (563) Google Scholar), with a notable exception of LysRS representatives, which belong to both classes (4Ibba M. Morgan S. Curnow A.W. Pridmore D.R. Vothknecht U.C. Gardner W. Lin W. Woese C.R. Söll D. Science. 1997; 278: 1119-1122Crossref PubMed Scopus (174) Google Scholar). Although the catalytic mechanisms of various aaRSs are broadly similar (5Zhang C.M. Perona J.J. Ryu K. Francklyn C. Hou Y.M. J. Mol. Biol. 2006; 361: 300-311Crossref PubMed Scopus (85) Google Scholar), each enzyme has developed a high specificity in recognizing its cognate amino acid and tRNA, which is pivotal for accurate translation of the genetic code (1Ibba M. Soll D. Annu. Rev. Biochem. 2000; 69: 617-650Crossref PubMed Scopus (1076) Google Scholar). The discrimination of the amino acids is based on recognizing the differences in the size and charge of the molecules (6Ataide S.F. Ibba M. ACS Chem. Biol. 2006; 1: 285-297Crossref PubMed Scopus (38) Google Scholar). The specificity of tRNA selection depends on a set of identity determinants that are mostly located at two distal extremities: the anticodon loop and the amino acid accepting stem. In a few instances, identity elements are also found in the D-arm, T-arm, and variable loop. They can either act as positive determinants that enhance aminoacylation or negative ones that prevent aminoacylation. The recognition of tRNAs by synthetases can also be affected by the modification of particular nucleotides (7Giegé R. Puglisi J.D. Florentz C. Prog. Nucleic Acid Res. Mol. Biol. 1993; 45: 129-206Crossref PubMed Scopus (218) Google Scholar, 8Giegé R. Sissler M. Florentz C. Nucleic Acids Res. 1998; 26: 5017-5035Crossref PubMed Scopus (628) Google Scholar). AaRSs show divergent strategies for tRNA recognition. Most notably, class I and class II aaRSs (including pyrrolysyl-tRNA synthetase, see Ref. 9Nozawa K. O'Donoghue P. Gundllapalli S. Araiso Y. Ishitani R. Umehara T. Söll D. Nureki O. Nature. 2009; 457: 1163-1167Crossref PubMed Scopus (126) Google Scholar) approach tRNAs from the minor and major groove sides of the acceptor stem, respectively (10Ribas de Pouplana L. Schimmel P. Cell. 2001; 104: 191-193Abstract Full Text Full Text PDF PubMed Scopus (142) Google Scholar). Although the majority of determinants are in direct contact with cognate synthetases (8Giegé R. Sissler M. Florentz C. Nucleic Acids Res. 1998; 26: 5017-5035Crossref PubMed Scopus (628) Google Scholar), the aminoacylation fidelity is controlled by kinetic differences more than by binding affinities (11Ebel J.P. Giegé R. Bonnet J. Kern D. Befort N. Bollack C. Fasiolo F. Gangloff J. Dirheimer G. Biochimie. 1973; 55: 547-557Crossref PubMed Scopus (154) Google Scholar). aminoacyl-tRNA synthetase (standard amino acid abbreviations precede RS throughout) seryl-tRNA synthetase methanogenic-type Methanosarcina barkeri SerRS M. barkeri tRNASer with anticodon GGA surface plasmon resonance wild type dithiothreitol 2-(N-morpholino)ethanesulfonic acid. aminoacyl-tRNA synthetase (standard amino acid abbreviations precede RS throughout) seryl-tRNA synthetase methanogenic-type Methanosarcina barkeri SerRS M. barkeri tRNASer with anticodon GGA surface plasmon resonance wild type dithiothreitol 2-(N-morpholino)ethanesulfonic acid. Seryl-tRNA synthetases (SerRSs), which catalyze the aminoacylation of several tRNASer isoacceptors and tRNASec with serine, can be divided into two structurally different groups: bacterial-type SerRSs function in a variety of archaeal, bacterial, and eukaryotic organisms, whereas the methanogenic-type was found only in methanogenic archaea (12Kim H.S. Vothknecht U.C. Hedderich R. Celic I. Söll D. J. Bacteriol. 1998; 180: 6446-6449Crossref PubMed Google Scholar, 13Tumbula D. Vothknecht U.C. Kim H.S. Ibba M. Min B. Li T. Pelaschier J. Stathopoulos C. Becker H. Söll D. Genetics. 1999; 152: 1269-1276PubMed Google Scholar). Furthermore, based on sequence comparison (14Lenhard B. Orellana O. Ibba M. Weygand-Duraseviæ I. Nucleic Acids Res. 1999; 27: 721-729Crossref PubMed Scopus (61) Google Scholar, 15Woese C.R. Olsen G.J. Ibba M. Söll D. Microbiol. Mol. Biol. Rev. 2000; 64: 202-236Crossref PubMed Scopus (530) Google Scholar) and x-ray analyses, two subgroups of bacterial-type SerRSs were identified: one consists of the enzymes from bacterial sources, best represented by those from Thermus thermophilus (16Fujinaga M. Berthet-Colominas C. Yaremchuk A.D. Tukalo M.A. Cusack S. J. Mol. Biol. 1993; 234: 222-233Crossref PubMed Scopus (92) Google Scholar) and Escherichia coli (3Cusack S. Berthet-Colominas C. Härtlein M. Nassar N. Leberman R. Nature. 1990; 347: 249-255Crossref PubMed Scopus (563) Google Scholar), and an archaeal/eukaryal-type, structurally related to SerRS from archaeon Pyrococcus horikoshii (17Itoh Y. Sekine S. Kuroishi C. Terada T. Shirouzu M. Kuramitsu S. Yokoyama S. RNA Biol. 2008; 5: 169-177Crossref PubMed Scopus (23) Google Scholar). All SerRSs are functional homodimers with a C-terminal active site domain typical for class II aaRSs and an N-terminal domain that is responsible for binding of the long variable arm of tRNASer isoacceptors (reviewed in Ref. 18Weygand-Durasevic I. Cusack S. The Aminoacyl-tRNA Synthetases.in: Ibba M. Francklyn C. Cusack S. Landes Bioscience, Georgetown, TX2005: 177-192Google Scholar), with exception of the mammalian mitochondrial enzyme (19Chimnaronk S. Gravers Jeppesen M. Suzuki T. Nyborg J. Watanabe K. EMBO J. 2005; 24: 3369-3379Crossref PubMed Scopus (70) Google Scholar). The long variable arm of tRNASer categorizes it as one of the type 2 tRNAs, including the tRNASer and tRNALeu species (from all organisms or domains of life) and bacterial tRNATyr species (20Sprinzl M. Vassilenko K.S. Nucleic Acids Res. 2005; 33: D139-D140Crossref PubMed Scopus (349) Google Scholar). In bacterial-type SerRS, the N-terminal domain forms an antiparallel α-helical coiled-coil structure (3Cusack S. Berthet-Colominas C. Härtlein M. Nassar N. Leberman R. Nature. 1990; 347: 249-255Crossref PubMed Scopus (563) Google Scholar, 16Fujinaga M. Berthet-Colominas C. Yaremchuk A.D. Tukalo M.A. Cusack S. J. Mol. Biol. 1993; 234: 222-233Crossref PubMed Scopus (92) Google Scholar), whereas in the methanogenic-type counterpart it is significantly larger and composed of a six-stranded antiparallel β-sheet capped by a bundle of three helices (H1, H2, and H4) with up-down topology and an additional short helix (H3) that runs almost perpendicular to helix H4 (21Bilokapic S. Maier T. Ahel D. Gruic-Sovulj I. Söll D. Weygand- Durasevic I. Ban N. EMBO J. 2006; 25: 2498-2509Crossref PubMed Scopus (72) Google Scholar) (see Fig. 1). Despite pronounced structural differences between the tRNA-binding domains in two SerRS types, in each case the N-terminal domain of one subunit interacts with the extra arm of tRNASer, to position the 3′-end of tRNA into the C-terminal active site of another subunit (23Biou V. Yaremchuk A. Tukalo M. Cusack S. Science. 1994; 263: 1404-1410Crossref PubMed Scopus (422) Google Scholar, 24Vincent C. Borel F. Willison J.C. Leberman R. Härtlein M. Nucleic Acids Res. 1995; 23: 1113-1118Crossref PubMed Scopus (36) Google Scholar, 25Bilokapic S. Ivic N. Godinic-Mikulcic V. Piantanida I. Ban N. Weygand-Durasevic I. J. Biol. Chem. 2009; 284: 10706-10713Abstract Full Text Full Text PDF PubMed Scopus (12) Google Scholar). The recent crystal structure of the first archaeal/eukaryal SerRS from the archaeon P. horikoshii, and the structure-based model of the enzyme bound with the T. thermophilus and P. horikoshii tRNAsSer, suggested that the helical N-terminal domain of P. horikoshii SerRS is also involved in the binding of the extra arm of tRNA (17Itoh Y. Sekine S. Kuroishi C. Terada T. Shirouzu M. Kuramitsu S. Yokoyama S. RNA Biol. 2008; 5: 169-177Crossref PubMed Scopus (23) Google Scholar). The recognition of tRNA by SerRS relies, besides on the long extra arm, on the identity elements in tRNASer acceptor arms, achieved by the motif 2 residues. Unlike the majority of aaRSs systems, the anticodon triplet is not recognized by SerRS. The first four base pairs in the tRNA acceptor arm (G1:C72, G2:C71, A/U3:U/A70, and R4:Y69) are identity elements for bacterial SerRS, and among them, the second G2:C71 base pair is the most significant (reviewed in Ref. 18Weygand-Durasevic I. Cusack S. The Aminoacyl-tRNA Synthetases.in: Ibba M. Francklyn C. Cusack S. Landes Bioscience, Georgetown, TX2005: 177-192Google Scholar). Consistently, the crystal structure of the T. thermophilus SerRS·tRNASer complex revealed that SerRS interacts with the tRNASer acceptor stem, and Ser261 is responsible for the base-specific interaction with G2 (23Biou V. Yaremchuk A. Tukalo M. Cusack S. Science. 1994; 263: 1404-1410Crossref PubMed Scopus (422) Google Scholar, 26Cusack S. Yaremchuk A. Tukalo M. EMBO J. 1996; 15: 2834-2842Crossref PubMed Scopus (155) Google Scholar). Although the acceptor stem sequences are not well conserved among the eukaryal tRNASer isoacceptors, the discriminator base G73 is an essential identity requirement for human tRNASer and serves as an anti-determinant in lower eukaryotes (reviewed in Ref. 18Weygand-Durasevic I. Cusack S. The Aminoacyl-tRNA Synthetases.in: Ibba M. Francklyn C. Cusack S. Landes Bioscience, Georgetown, TX2005: 177-192Google Scholar). Unlike eukaryal tRNAsSer, archaeal tRNAsSer conserve the G1:C72, C2:G71, C3:G70, and G4:C69 base pairs in the acceptor stem. However, the tRNA specificity of the archaeal/eukaryal SerRS from P. horikoshii seems to depend mainly on the extra arm, but not on the acceptor stem. Indeed, this enzyme exhibits quite relaxed specificity for tRNASer recognition (17Itoh Y. Sekine S. Kuroishi C. Terada T. Shirouzu M. Kuramitsu S. Yokoyama S. RNA Biol. 2008; 5: 169-177Crossref PubMed Scopus (23) Google Scholar). Archaeon Methanosarcina barkeri, which possesses two dissimilar SerRSs, one of a bacterial- and the other of methanogenic-type, provides an excellent system for studying the evolution of tRNASer determinants. Two enzymes recognize the same set of tRNA isoacceptors in vitro (27Korencic D. Polycarpo C. Weygand-Durasevic I. Söll D. J. Biol. Chem. 2004; 279: 48780-48786Abstract Full Text Full Text PDF PubMed Scopus (35) Google Scholar). We have undertaken several approaches to elucidate the basis for their different serylation mechanisms. Kinetic analysis of variant tRNASer transcripts by the two archaeal SerRS enzymes (27Korencic D. Polycarpo C. Weygand-Durasevic I. Söll D. J. Biol. Chem. 2004; 279: 48780-48786Abstract Full Text Full Text PDF PubMed Scopus (35) Google Scholar) revealed that the length of the variable arm is a critical recognition element for both enzymes, as is the identity of the discriminator base (G73) and base pair G30:C40 in the anticodon stem. However, additional determinants were identified as being required for specific serylation by the unusual methanogenic-type enzyme, which relies on G1:C72 identity and on the number of unpaired nucleotides at the base of the variable loop. The tRNA recognition pattern by two M. barkeri SerRS may differ in vivo, because only bacterial-type SerRS complements the function of thermolabile E. coli SerRS (28Lesjak S. Weygand-Durasevic I. FEMS Microbiol. Lett. 2009; 294: 111-118Crossref PubMed Scopus (7) Google Scholar). We are grateful to Eilika Weber-Ban for useful suggestions regarding experimental procedures and for valuable discussions. Mike Scott and Stefan Schauer from the Functional Genomics Center, Zurich, are greatly acknowledged for assistance in SPR experiments. We are indebted to Rouven Bingel-Erlenmeyer and Martina Trokter for help with protein purification. Supplementary Material Download .pdf (.03 MB) Help with pdf files Download .pdf (.03 MB) Help with pdf files
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