Subsystem: YrdC-YciO-Sua5 protein family
This subsystem's description is:
This subsystem has been originally encoded by Dr. El Yacoubi, whose contribution to the SEED database we gratefully acknowledge.
Plant-Prokaryote project SUMMARY for YrdC-Sua5 family (At5g60590 in Arabidopsis):
Using a combination of bioinformatic, genetic, structural and biochemical approaches, the universal protein family YrdC/Sua5 (COG0009) was shown to be involved in the biosynthesis of the N6-threonylcarbamoyl adenosine (t6A) – an anticodon-loop modification found at position 37 of tRNAs decoding ANN codons (El Yacoubi et al., 2009).
OBSERVATIONS/BACKGROUND:
N6-threonylcarbamoyl adenosine (t6A) is a universal modification found at position 37 of ANN decoding tRNAs, which imparts a unique structure to the anticodon loop enhancing its binding to ribosomes in vitro. Although considerable biochemical and biophysical information exists on the function of this hypermodified base (Elkins et al., 1974; Yarian et al., 2002; Durant et al., 2005), the t6A biosynthesis pathway has only been partially biochemically characterized and shown to be an ATP-dependent process requiring threonine and carbonate although none of the genes involved in its biosynthesis have been identified.
1.Analysis of all sequenced tRNAs (38) suggested that all organisms should have a t6A biosynthesis pathway.
2.The list of orthologous protein families shared between Gram(+) and Gram(-) organisms, including those with severely reduced genomes, yielded only 95 “universal” families, mostly related to translation (ribosomal protein, tRNA synthetases and tRNA modification).
3. From this list, the YrdC/Sua5 family (assigned to COG009) was the most probable candidate for an enzyme involved in t6A biosynthesis for the following reasons: (i) all organisms sequenced to date have a homolog of the YrdC/Sua5 proteins, some organisms, such as E. coli, have two representatives (YrdC and YciO), but most have only one; (ii) the YrdC domain is also found in the enzyme family HypF [involved in the maturation of the metal center of the Ni-Fe hydrogenase HypE] which catalyzes chemistries similar to the ones expected in t6A biosynthesis (Elkin et al., 1974); (iii) YrdC and YciO of E. coli were found to bind double-stranded RNA and tRNA (Teplova et al., 2000); (iv) Sua5 in yeast was proposed to be involved in translation (Na et al., 1992).
3. We performed LC–MS/MS analysis of yeast tRNA extracted from different yeast strains, which linked the disappearance of the t6A peak to the deletion of sua5 (El Yacoubi, et al., 2009).
4. Structural and biochemical analyses revealed that the E. coli YrdC protein binds ATP and preferentially binds RNA(Thr) lacking only the t6A modification (El Yacoubi, et al., 2009).
5. We also investigated the essentiality of this gene family in two biological systems, E. coli and yeast and proved that yrdC is essential in E. coli, whereas SUA5 is dispensable in yeast but results in severe growth phenotypes (El Yacoubi, et al., 2009).
6. However, specific biochemical function for the YrdC/Sua5 proteins (i.e. catalytic activity) aside from tRNA and ATP binding (El Yacoubi, et al., 2009) and ATPase activity (Agari et al., 2008), has not been determined. Were unable to show threonine binding by YrdC, under ATP binding conditions, nor incorporation of threonine in a tRNAThr transcript in the presence of YrdC. Further experimentation is needed to determine the role of YrdC in t6A biosynthesis.
7. Although iso-functional, the YrdC and Sua5 subfamilies are distinguished in this database based their domain structure: while both subfamilies contain the YrdC domain, only the Sua5 proteins carry an additional C-terminal Sua5 domain
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Plant-Prokaryote DOE project SUMMARY for YciO and YciV families (in Arabidopsis: At3g01920 and At2g13840 respectively)
Most organisms harbor only one member of the YrdC/Sua5 (COG009) protein family. Exceptions include plants and several bacteria that have two homologs. Expression of the second COG009 encoding gene of E. coli, yciO, failed to complement the t6A-minus phenotype of yeast strains lacking a functional SUA5 gene (El Yacoubi, et al., 2009). The two subfamilies can also be distinguished by sequence analysis, as residues K50 and R52 are strictly conserved in the YrdC/Sua5 subfamily but absent in YciO sequences. These residues are essential for YrdC/Sua5 function as shown by site directed mutagenesis and complementation experiments.
Therefore the YciO branch of COG009 is separated in this database from the YrdC and Sua5 subfamilies, as the results presented here suggest that function of the YciO subgroup of COG0009 is distinct from that of YrdC/Sua5 (El Yacoubi, et al., 2009). It is still unknown.
Protein family YciV appears to be involved in strong functional coupling with the YciO subfamily (as the corresponding genes appear to co-occur and co-localize in the majority of genomes where they are present). We predict that the YciO and YciV protein families are jointly involved in yet unknown modification of RNA or proteins
REFERENCES
1. El Yacoubi B, Lyons B, Cruz Y, Reddy R, Nordin B, Agnelli F, Williamson JR, Schimmel P, Swairjo MA, de Crécy-Lagard V. 2009. The universal YrdC/Sua5 family is required for the formation of threonylcarbamoyladenosine in tRNA. Nucleic Acids Res, 37(9):2894-909.
Agari Y, Sato S, Wakamatsu T, Bessho Y, Ebihara A, Yokoyama S, Kuramitsu S, Shinkai A. X-ray crystal structure of a hypothetical Sua5 protein from Sulfolobus tokodaii strain. Proteins (2008) 70:1108–1111
Durant PC, Bajji AC, Sundaram M, Kumar RK, Davis DR. Structural effects of hypermodified nucleosides in the Escherichia coli and human tRNALys anticodon loop: the effect of nucleosides s(2)U, mcm(5)U, mcm(5)s(2)U, mnm(5)s(2)U, t(6)A, and ms(2)t(6)A. Biochemistry (2005) 44:8078–8089.
Elkins BN, Keller EB. The enzymatic synthesis of N-(purin-6-ylcarbamoyl)threonine, an anticodon-adjacent base in transfer ribonucleic acid. Biochemistry (1974) 13:4622–4628
Na JG, Pinto I, Hampsey M. Isolation and characterization of SUA5, a novel gene required for normal growth in Saccharomyces cerevisiae. Genetics (1992) 131:791–801.
Teplova M, Tereshko V, Sanishvili R, Joachimiak A, Bushueva T, Anderson WF, Egli M. The structure of the yrdC gene product from Escherichia coli reveals a new fold and suggests a role in RNA binding. Protein Sci. (2000) 9:2557–2566.
Yarian C, Townsend H, Czestkowski W, Sochacka E, Malkiewicz AJ, Guenther R, Miskiewicz A, Agris PF. Accurate translation of the genetic code depends on tRNA modified nucleosides. J. Biol. Chem. (2002) 277:16391–16395
For more information, please check out the description and the additional notes tabs, below
| Diagram | Functional Roles | Subsystem Spreadsheet | Description | Additional Notes | |||||||||
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This subsystem has been originally encoded by Dr. El Yacoubi, whose contribution to the SEED database we gratefully acknowledge. Plant-Prokaryote project SUMMARY for YrdC-Sua5 family (At5g60590 in Arabidopsis): Using a combination of bioinformatic, genetic, structural and biochemical approaches, the universal protein family YrdC/Sua5 (COG0009) was shown to be involved in the biosynthesis of the N6-threonylcarbamoyl adenosine (t6A) – an anticodon-loop modification found at position 37 of tRNAs decoding ANN codons (El Yacoubi et al., 2009). OBSERVATIONS/BACKGROUND: N6-threonylcarbamoyl adenosine (t6A) is a universal modification found at position 37 of ANN decoding tRNAs, which imparts a unique structure to the anticodon loop enhancing its binding to ribosomes in vitro. Although considerable biochemical and biophysical information exists on the function of this hypermodified base (Elkins et al., 1974; Yarian et al., 2002; Durant et al., 2005), the t6A biosynthesis pathway has only been partially biochemically characterized and shown to be an ATP-dependent process requiring threonine and carbonate although none of the genes involved in its biosynthesis have been identified. 1.Analysis of all sequenced tRNAs (38) suggested that all organisms should have a t6A biosynthesis pathway. 2.The list of orthologous protein families shared between Gram(+) and Gram(-) organisms, including those with severely reduced genomes, yielded only 95 “universal” families, mostly related to translation (ribosomal protein, tRNA synthetases and tRNA modification). 3. From this list, the YrdC/Sua5 family (assigned to COG009) was the most probable candidate for an enzyme involved in t6A biosynthesis for the following reasons: (i) all organisms sequenced to date have a homolog of the YrdC/Sua5 proteins, some organisms, such as E. coli, have two representatives (YrdC and YciO), but most have only one; (ii) the YrdC domain is also found in the enzyme family HypF [involved in the maturation of the metal center of the Ni-Fe hydrogenase HypE] which catalyzes chemistries similar to the ones expected in t6A biosynthesis (Elkin et al., 1974); (iii) YrdC and YciO of E. coli were found to bind double-stranded RNA and tRNA (Teplova et al., 2000); (iv) Sua5 in yeast was proposed to be involved in translation (Na et al., 1992). 3. We performed LC–MS/MS analysis of yeast tRNA extracted from different yeast strains, which linked the disappearance of the t6A peak to the deletion of sua5 (El Yacoubi, et al., 2009). 4. Structural and biochemical analyses revealed that the E. coli YrdC protein binds ATP and preferentially binds RNA(Thr) lacking only the t6A modification (El Yacoubi, et al., 2009). 5. We also investigated the essentiality of this gene family in two biological systems, E. coli and yeast and proved that yrdC is essential in E. coli, whereas SUA5 is dispensable in yeast but results in severe growth phenotypes (El Yacoubi, et al., 2009). 6. However, specific biochemical function for the YrdC/Sua5 proteins (i.e. catalytic activity) aside from tRNA and ATP binding (El Yacoubi, et al., 2009) and ATPase activity (Agari et al., 2008), has not been determined. Were unable to show threonine binding by YrdC, under ATP binding conditions, nor incorporation of threonine in a tRNAThr transcript in the presence of YrdC. Further experimentation is needed to determine the role of YrdC in t6A biosynthesis. 7. Although iso-functional, the YrdC and Sua5 subfamilies are distinguished in this database based their domain structure: while both subfamilies contain the YrdC domain, only the Sua5 proteins carry an additional C-terminal Sua5 domain * * * * * * * * * * * * * * * * * * * * Plant-Prokaryote DOE project SUMMARY for YciO and YciV families (in Arabidopsis: At3g01920 and At2g13840 respectively) Most organisms harbor only one member of the YrdC/Sua5 (COG009) protein family. Exceptions include plants and several bacteria that have two homologs. Expression of the second COG009 encoding gene of E. coli, yciO, failed to complement the t6A-minus phenotype of yeast strains lacking a functional SUA5 gene (El Yacoubi, et al., 2009). The two subfamilies can also be distinguished by sequence analysis, as residues K50 and R52 are strictly conserved in the YrdC/Sua5 subfamily but absent in YciO sequences. These residues are essential for YrdC/Sua5 function as shown by site directed mutagenesis and complementation experiments. Therefore the YciO branch of COG009 is separated in this database from the YrdC and Sua5 subfamilies, as the results presented here suggest that function of the YciO subgroup of COG0009 is distinct from that of YrdC/Sua5 (El Yacoubi, et al., 2009). It is still unknown. Protein family YciV appears to be involved in strong functional coupling with the YciO subfamily (as the corresponding genes appear to co-occur and co-localize in the majority of genomes where they are present). We predict that the YciO and YciV protein families are jointly involved in yet unknown modification of RNA or proteins REFERENCES 1. El Yacoubi B, Lyons B, Cruz Y, Reddy R, Nordin B, Agnelli F, Williamson JR, Schimmel P, Swairjo MA, de Crécy-Lagard V. 2009. The universal YrdC/Sua5 family is required for the formation of threonylcarbamoyladenosine in tRNA. Nucleic Acids Res, 37(9):2894-909. Agari Y, Sato S, Wakamatsu T, Bessho Y, Ebihara A, Yokoyama S, Kuramitsu S, Shinkai A. X-ray crystal structure of a hypothetical Sua5 protein from Sulfolobus tokodaii strain. Proteins (2008) 70:1108–1111 Durant PC, Bajji AC, Sundaram M, Kumar RK, Davis DR. Structural effects of hypermodified nucleosides in the Escherichia coli and human tRNALys anticodon loop: the effect of nucleosides s(2)U, mcm(5)U, mcm(5)s(2)U, mnm(5)s(2)U, t(6)A, and ms(2)t(6)A. Biochemistry (2005) 44:8078–8089. Elkins BN, Keller EB. The enzymatic synthesis of N-(purin-6-ylcarbamoyl)threonine, an anticodon-adjacent base in transfer ribonucleic acid. Biochemistry (1974) 13:4622–4628 Na JG, Pinto I, Hampsey M. Isolation and characterization of SUA5, a novel gene required for normal growth in Saccharomyces cerevisiae. Genetics (1992) 131:791–801. Teplova M, Tereshko V, Sanishvili R, Joachimiak A, Bushueva T, Anderson WF, Egli M. The structure of the yrdC gene product from Escherichia coli reveals a new fold and suggests a role in RNA binding. Protein Sci. (2000) 9:2557–2566. Yarian C, Townsend H, Czestkowski W, Sochacka E, Malkiewicz AJ, Guenther R, Miskiewicz A, Agris PF. Accurate translation of the genetic code depends on tRNA modified nucleosides. J. Biol. Chem. (2002) 277:16391–16395 Plant-Prokaryote project SUMMARY for YgjD/Kae1 family (At2g45270, At4g22720 in Arabidopsis): Based on a comparative genomic and structural analysis we predicted this family was involved in the biosynthesis of N6-threonylcarbamoyl adenosine, a universal modification found at position 37 of tRNAs decoding ANN codons (El Yacoubi et al., submitted). This was confirmed as a yeast mutant lacking Kae1 is devoid of t6A. The t6A-deletant strains were also used to reveal that t6A has a critical role in initiation codon restriction to AUG and in restricting frameshifiting at tandem ANN codons (El Yacoubi et al., submitted). OBSERVATIONS/BACKGROUND: The YgjD/Kae1 family (COG0533) has been on the top-ten list of universally conserved proteins of unknown function for over five years (Galperin & Koonin, 2004). It is essential in Escherichia coli, Bacillus subtilis, and several other bacteria where examined (Arigoni et al., 1998; Zalacain et al., 2003). It has been linked to DNA maintenance in bacteria and mitochondria and transcription regulation and telomere homeostasis in eukaryotes (Downey et al., 2006; Kisseleva-Romanova et al., 2006), but its actual function has never been found (Hecker et al., 2009). Early work on the Pasteurella haemolytica homolog has erroneously assigned this family the function of O-sialoglycoprotein peptidase (Abdullah et al., 1991; Abdullah et al., 1992). Furthermore, it became the founding member of the M22 class of proteases, aggravating this mistake. No other member of the protein family has this activity demonstrated, and even the Pasteurella protein expressed in E. coli lacks the activity. In Archaea the YgjD proteins are often fused with p53-regulating protein kinase [PRPK (human) // YGR262c (yeast)]. The human protein is able to partially suppress the effects of a yeast mutation (the phenotype of which is abnormal budding). The yeast protein is able to phosphorylate human p53. For this reason the PRPK // YGR262c family is included in this SS * * * * * * * * * * * * * * * * Plant-Prokaryote project SUMMARY for YaeZ family (no eukaryotic homologs): We also showed that YaeZ, a YgjD paralog (Nichols et al., 2006), is required for YgjD function in vivo in bacteria. Indeed the Bacillus subtilis ortholog of ygjD gene, ydiE, cannot complement the essentiality of the Escherichia coli ygjD gene in the absence of the B. subtilis yeaZ ortholog ydiC (El Yacoubi B. et al., submitted) Similar to the YgjD family, YaeZ orthologs are also nearly universal among Bacteria, but missing from Eukarya and Archaea. YeaZ is essential in Escherichia coli K12 (Handford et al., 2009). Conditional expression strains for YaeZ, YgjD, and YjeE genes in EC show dramatic changes in cell ultrastructure (Handford et al., 2009). Repression of the expression of yeaZ results in cells with highly condensed nucleoids, while repression of yjeE and ygjD expression results in at least a proportion of very enlarged cells with an unusual peripheral distribution of DNA. The results of bacterial two-hybrid experiments show that YeaZ can interact with both YjeE and YgjD but that YgjD is the preferred interaction partner. The results of in vitro experiments indicate that YeaZ mediates the proteolysis of YgjD, suggesting that YeaZ and YjeE act as regulators to control the activity of this protein. Our results are consistent with these proteins forming a link between DNA metabolism and cell division (Handford et al., 2009) REFERENCES: 1. El Yacoubi B. et al., 2010 submitted Abdullah KM, Lo RY, Mellors A. 1991. Cloning, nucleotide sequence, and expression of the Pasteurella haemolytica A1 glycoprotease gene. J. Bacteriol. 173(18):5597-603. PMID: 1885539 Abdullah KM, Udoh EA, Shewen PE, Mellors A. 1992. A neutral glycoprotease of Pasteurella haemolytica A1 specifically cleaves O-sialoglycoproteins. Infect. Immun. 60(1):56-62. PMID: 1729196 Arigoni F, Talabot F, Peitsch M, Edgerton MD, Meldrum E, Allet E, Fish R, Jamotte T, Curchod ML, Loferer H. 1998. A genome-based approach for the identification of essential bacterial genes. Nat. Biotechnol. 16(9):851-6. PMID: 9743119 Downey,M., Houlsworth,R., Maringele,L., et al. (2006) A genome-wide screen identifies the evolutionarily conserved KEOPS complex as a telomere regulator. Cell, 124, 1155–1168. Galperin,M.Y. and Koonin,E.V. (2004) ‘Conserved hypothetical’ proteins: prioritization of targets for experimental study. Nucleic Acids Res., 32, 5452–5463. Handford JI, Ize B, Buchanan G, Butland GP, Greenblatt J, Emili A, Palmer T. 2009. Conserved network of proteins essential for bacterial viability. J Bacteriol, 191(15):4732-49 Hecker A, Leulliot N, Gadelle D, Graille M, Justome A, Dorlet P, Brochier C, Quevillon-Cheruel S, Le Cam E, van Tilbeurgh H, Forterre P. 2007. An archaeal orthologue of the universal protein Kae1 is an iron metalloprotein which exhibits atypical DNA-binding properties and apurinic-endonuclease activity in vitro. Nucleic Acids Res, 35(18):6042-51 Kisseleva-Romanova,E., Lopreiato,R., Baudin-Baillieu,A., Rousselle,J.C., Ilan,L., Hofmann,K., Namane,A., Mann,C. and Libri,D. (2006) Yeast homolog of a cancer-testis antigen defines a new transcription complex. EMBO J., 25, 3576–3585. Nichols CE, Johnson C, Lockyer M, Charles IG, Lamb HK, Hawkins AR, Stammers DK. 2006. Structural characterization of Salmonella typhimurium YeaZ, an M22 O-sialoglycoprotein endopeptidase homolog. Proteins 64(1):111-23. PMID: 16617437 Zalacain M, Biswas S, Ingraham KA, Ambrad J, Bryant A, Chalker AF, Iordanescu S, Fan J, Fan F, Lunsford RD, O'Dwyer K, Palmer LM, So C, Sylvester D, Volker C, Warren P, McDevitt D, Brown JR, Holmes DJ, Burnham MK. 2003. A global approach to identify novel broad-spectrum antibacterial targets among proteins of unknown function. J. Mol. Microbiol. Biotechnol. 6(2):109-26. PMID: 15044829 | |||||||||||||
