Proteopedia

5m04

Structure of ObgE from Escherichia coliStructure of ObgE from Escherichia coli

Structural highlights

5m04 is a 1 chain structure with sequence from Escherichia coli DH5alpha. Full crystallographic information is available from OCA. For a guided tour on the structure components use FirstGlance.
Method:X-ray diffraction, Resolution 1.85Å
Ligands:,
Resources:FirstGlance, OCA, PDBe, RCSB, PDBsum, ProSAT

Function

OBG_ECOLI An abundant, essential GTPase which binds GTP, GDP and ppGpp with moderate affinity, shows high guanine nucleotide exchange rate constants for GTP and GDP, and has a relatively low GTP hydrolysis rate. It is estimated there are 34000 molecules in log-phase cells and 5600 molecules in stationary-phase cells as the monomer. Required for chromosome segregation. It also plays a role in the stringent response, and a non-essential role in the late steps of ribosome biogenesis, but how it performs these functions remains unelucidated.[1] [2] [3] [4] [5] [6] [7] [8] [9] [10] [11] [12] Required for correct chromosome partitioning; in temperature-sensitive (ts) mutant nucleoids do not partition but remain in the middle of cell, cells elongate but do not divide. Overexpression protects cells against UV damage. Ts mutants have impaired plasmid and lamdba phage replication, possibly via effects on DnaA. Regulates DnaA levels. Genetic interactions of Val-168 and a C-terminal insertion mutant with the double-strand break repair factors recA and recBCD, and with seqA suggests that ObgE, either directly or indirectly, promotes replication fork stability. Initiation of DNA replication continues in ObgE-depleted cells.[13] [14] [15] [16] [17] [18] [19] [20] [21] [22] [23] [24] Binds to pre-50S ribosomal subunits in a salt-dependent manner, has also been seen associated with 30S subunits. Overexpression rescues an rrmJ deletion stabilizing the 70S ribosome. Even at permissive temperatures the ts mutant (Gln-80/Asn-85) shows disrupted 50S ribosomal subunit assembly, defects in 16 and 23S rRNA processing and altered association of some late-assembling ribosomal proteins. Dissociates from the pre-50S ribosome under conditions of amino acid starvation. The levels of (p)ppGpp rise in the ts mutant (Gln-80/Asn-85), possibly because ObgE controls SpoT. Binds GDP and ppGpp with the same affinity. During ribosome assembly ObgE acts later than the rRNA methyltransferase rrmj and DEAD-box RNA helicases DeaD and SrmB.[25] [26] [27] [28] [29] [30] [31] [32] [33] [34] [35] [36]

Publication Abstract from PubMed

The Obg protein family belongs to the TRAFAC (translation factor) class of P-loop GTPases and is conserved from bacteria to eukaryotes. Essential roles in many different cellular processes have been suggested for the Obg protein from Escherichia coli (ObgE), and we recently showed that it is a central regulator of bacterial persistence. Here, we report the first crystal structure of ObgE at 1.85 A resolution in the GDP-bound state, showing the characteristic N-terminal domain and a central G domain that are common to all Obg proteins. ObgE also contains an intrinsically disordered C-terminal domain, and we show here that this domain specifically contributed to GTP binding, while it did not influence GDP binding or GTP hydrolysis. Biophysical analysis, using small angle X-ray scattering and multi-angle light scattering experiments, revealed that ObgE is a monomer in solution, regardless of the bound nucleotide. In contrast to recent suggestions, our biochemical analyses further indicate that ObgE is neither activated by K+ ions nor by homodimerization. However, the ObgE GTPase activity was stimulated upon binding to the ribosome, confirming the ribosome-dependent GTPase activity of the Obg family. Combined, our data represent an important step toward further unraveling the detailed molecular mechanism of ObgE, which might pave the way to further studies into how this GTPase regulates bacterial physiology, including persistence.

Structural and Biochemical Analysis of Escherichia coli ObgE, a Central Regulator of Bacterial Persistence.,Gkekas S, Singh RK, Shkumatov AV, Messens J, Fauvart M, Verstraeten N, Michiels J, Versees W J Biol Chem. 2017 Feb 21. pii: jbc.M116.761809. doi: 10.1074/jbc.M116.761809. PMID:28223358[37]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.

See Also

References

  1. ↑ Kobayashi G, Moriya S, Wada C. Deficiency of essential GTP-binding protein ObgE in Escherichia coli inhibits chromosome partition. Mol Microbiol. 2001 Sep;41(5):1037-51. PMID:11555285
  2. ↑ Dutkiewicz R, Slominska M, Wegrzyn G, Czyz A. Overexpression of the cgtA (yhbZ, obgE) gene, coding for an essential GTP-binding protein, impairs the regulation of chromosomal functions in Escherichia coli. Curr Microbiol. 2002 Dec;45(6):440-5. PMID:12402086 doi:http://dx.doi.org/10.1007/s00284-002-3713-x
  3. ↑ Tan J, Jakob U, Bardwell JC. Overexpression of two different GTPases rescues a null mutation in a heat-induced rRNA methyltransferase. J Bacteriol. 2002 May;184(10):2692-8. PMID:11976298
  4. ↑ Zielke R, Sikora A, Dutkiewicz R, Wegrzyn G, Czyz A. Involvement of the cgtA gene function in stimulation of DNA repair in Escherichia coli and Vibrio harveyi. Microbiology. 2003 Jul;149(Pt 7):1763-70. PMID:12855728
  5. ↑ Ulanowska K, Sikora A, Wegrzyn G, Czyz A. Role of the cgtA gene function in DNA replication of extrachromosomal elements in Escherichia coli. Plasmid. 2003 Jul;50(1):45-52. PMID:12826057
  6. ↑ Sato A, Kobayashi G, Hayashi H, Yoshida H, Wada A, Maeda M, Hiraga S, Takeyasu K, Wada C. The GTP binding protein Obg homolog ObgE is involved in ribosome maturation. Genes Cells. 2005 May;10(5):393-408. PMID:15836769 doi:http://dx.doi.org/10.1111/j.1365-2443.2005.00851.x
  7. ↑ Foti JJ, Schienda J, Sutera VA Jr, Lovett ST. A bacterial G protein-mediated response to replication arrest. Mol Cell. 2005 Feb 18;17(4):549-60. PMID:15721258 doi:http://dx.doi.org/10.1016/j.molcel.2005.01.012
  8. ↑ Sikora AE, Zielke R, Wegrzyn A, Wegrzyn G. DNA replication defect in the Escherichia coli cgtA(ts) mutant arising from reduced DnaA levels. Arch Microbiol. 2006 Jun;185(5):340-7. Epub 2006 Mar 4. PMID:16518617 doi:http://dx.doi.org/10.1007/s00203-006-0099-3
  9. ↑ Jiang M, Datta K, Walker A, Strahler J, Bagamasbad P, Andrews PC, Maddock JR. The Escherichia coli GTPase CgtAE is involved in late steps of large ribosome assembly. J Bacteriol. 2006 Oct;188(19):6757-70. PMID:16980477 doi:http://dx.doi.org/10.1128/JB.00444-06
  10. ↑ Jiang M, Sullivan SM, Wout PK, Maddock JR. G-protein control of the ribosome-associated stress response protein SpoT. J Bacteriol. 2007 Sep;189(17):6140-7. Epub 2007 Jul 6. PMID:17616600 doi:http://dx.doi.org/10.1128/JB.00315-07
  11. ↑ Foti JJ, Persky NS, Ferullo DJ, Lovett ST. Chromosome segregation control by Escherichia coli ObgE GTPase. Mol Microbiol. 2007 Jul;65(2):569-81. Epub 2007 Jun 18. PMID:17578452 doi:http://dx.doi.org/10.1111/j.1365-2958.2007.05811.x
  12. ↑ Persky NS, Ferullo DJ, Cooper DL, Moore HR, Lovett ST. The ObgE/CgtA GTPase influences the stringent response to amino acid starvation in Escherichia coli. Mol Microbiol. 2009 Jul;73(2):253-66. doi: 10.1111/j.1365-2958.2009.06767.x. Epub, 2009 Jun 23. PMID:19555460 doi:http://dx.doi.org/10.1111/j.1365-2958.2009.06767.x
  13. ↑ Kobayashi G, Moriya S, Wada C. Deficiency of essential GTP-binding protein ObgE in Escherichia coli inhibits chromosome partition. Mol Microbiol. 2001 Sep;41(5):1037-51. PMID:11555285
  14. ↑ Dutkiewicz R, Slominska M, Wegrzyn G, Czyz A. Overexpression of the cgtA (yhbZ, obgE) gene, coding for an essential GTP-binding protein, impairs the regulation of chromosomal functions in Escherichia coli. Curr Microbiol. 2002 Dec;45(6):440-5. PMID:12402086 doi:http://dx.doi.org/10.1007/s00284-002-3713-x
  15. ↑ Tan J, Jakob U, Bardwell JC. Overexpression of two different GTPases rescues a null mutation in a heat-induced rRNA methyltransferase. J Bacteriol. 2002 May;184(10):2692-8. PMID:11976298
  16. ↑ Zielke R, Sikora A, Dutkiewicz R, Wegrzyn G, Czyz A. Involvement of the cgtA gene function in stimulation of DNA repair in Escherichia coli and Vibrio harveyi. Microbiology. 2003 Jul;149(Pt 7):1763-70. PMID:12855728
  17. ↑ Ulanowska K, Sikora A, Wegrzyn G, Czyz A. Role of the cgtA gene function in DNA replication of extrachromosomal elements in Escherichia coli. Plasmid. 2003 Jul;50(1):45-52. PMID:12826057
  18. ↑ Sato A, Kobayashi G, Hayashi H, Yoshida H, Wada A, Maeda M, Hiraga S, Takeyasu K, Wada C. The GTP binding protein Obg homolog ObgE is involved in ribosome maturation. Genes Cells. 2005 May;10(5):393-408. PMID:15836769 doi:http://dx.doi.org/10.1111/j.1365-2443.2005.00851.x
  19. ↑ Foti JJ, Schienda J, Sutera VA Jr, Lovett ST. A bacterial G protein-mediated response to replication arrest. Mol Cell. 2005 Feb 18;17(4):549-60. PMID:15721258 doi:http://dx.doi.org/10.1016/j.molcel.2005.01.012
  20. ↑ Sikora AE, Zielke R, Wegrzyn A, Wegrzyn G. DNA replication defect in the Escherichia coli cgtA(ts) mutant arising from reduced DnaA levels. Arch Microbiol. 2006 Jun;185(5):340-7. Epub 2006 Mar 4. PMID:16518617 doi:http://dx.doi.org/10.1007/s00203-006-0099-3
  21. ↑ Jiang M, Datta K, Walker A, Strahler J, Bagamasbad P, Andrews PC, Maddock JR. The Escherichia coli GTPase CgtAE is involved in late steps of large ribosome assembly. J Bacteriol. 2006 Oct;188(19):6757-70. PMID:16980477 doi:http://dx.doi.org/10.1128/JB.00444-06
  22. ↑ Jiang M, Sullivan SM, Wout PK, Maddock JR. G-protein control of the ribosome-associated stress response protein SpoT. J Bacteriol. 2007 Sep;189(17):6140-7. Epub 2007 Jul 6. PMID:17616600 doi:http://dx.doi.org/10.1128/JB.00315-07
  23. ↑ Foti JJ, Persky NS, Ferullo DJ, Lovett ST. Chromosome segregation control by Escherichia coli ObgE GTPase. Mol Microbiol. 2007 Jul;65(2):569-81. Epub 2007 Jun 18. PMID:17578452 doi:http://dx.doi.org/10.1111/j.1365-2958.2007.05811.x
  24. ↑ Persky NS, Ferullo DJ, Cooper DL, Moore HR, Lovett ST. The ObgE/CgtA GTPase influences the stringent response to amino acid starvation in Escherichia coli. Mol Microbiol. 2009 Jul;73(2):253-66. doi: 10.1111/j.1365-2958.2009.06767.x. Epub, 2009 Jun 23. PMID:19555460 doi:http://dx.doi.org/10.1111/j.1365-2958.2009.06767.x
  25. ↑ Kobayashi G, Moriya S, Wada C. Deficiency of essential GTP-binding protein ObgE in Escherichia coli inhibits chromosome partition. Mol Microbiol. 2001 Sep;41(5):1037-51. PMID:11555285
  26. ↑ Dutkiewicz R, Slominska M, Wegrzyn G, Czyz A. Overexpression of the cgtA (yhbZ, obgE) gene, coding for an essential GTP-binding protein, impairs the regulation of chromosomal functions in Escherichia coli. Curr Microbiol. 2002 Dec;45(6):440-5. PMID:12402086 doi:http://dx.doi.org/10.1007/s00284-002-3713-x
  27. ↑ Tan J, Jakob U, Bardwell JC. Overexpression of two different GTPases rescues a null mutation in a heat-induced rRNA methyltransferase. J Bacteriol. 2002 May;184(10):2692-8. PMID:11976298
  28. ↑ Zielke R, Sikora A, Dutkiewicz R, Wegrzyn G, Czyz A. Involvement of the cgtA gene function in stimulation of DNA repair in Escherichia coli and Vibrio harveyi. Microbiology. 2003 Jul;149(Pt 7):1763-70. PMID:12855728
  29. ↑ Ulanowska K, Sikora A, Wegrzyn G, Czyz A. Role of the cgtA gene function in DNA replication of extrachromosomal elements in Escherichia coli. Plasmid. 2003 Jul;50(1):45-52. PMID:12826057
  30. ↑ Sato A, Kobayashi G, Hayashi H, Yoshida H, Wada A, Maeda M, Hiraga S, Takeyasu K, Wada C. The GTP binding protein Obg homolog ObgE is involved in ribosome maturation. Genes Cells. 2005 May;10(5):393-408. PMID:15836769 doi:http://dx.doi.org/10.1111/j.1365-2443.2005.00851.x
  31. ↑ Foti JJ, Schienda J, Sutera VA Jr, Lovett ST. A bacterial G protein-mediated response to replication arrest. Mol Cell. 2005 Feb 18;17(4):549-60. PMID:15721258 doi:http://dx.doi.org/10.1016/j.molcel.2005.01.012
  32. ↑ Sikora AE, Zielke R, Wegrzyn A, Wegrzyn G. DNA replication defect in the Escherichia coli cgtA(ts) mutant arising from reduced DnaA levels. Arch Microbiol. 2006 Jun;185(5):340-7. Epub 2006 Mar 4. PMID:16518617 doi:http://dx.doi.org/10.1007/s00203-006-0099-3
  33. ↑ Jiang M, Datta K, Walker A, Strahler J, Bagamasbad P, Andrews PC, Maddock JR. The Escherichia coli GTPase CgtAE is involved in late steps of large ribosome assembly. J Bacteriol. 2006 Oct;188(19):6757-70. PMID:16980477 doi:http://dx.doi.org/10.1128/JB.00444-06
  34. ↑ Jiang M, Sullivan SM, Wout PK, Maddock JR. G-protein control of the ribosome-associated stress response protein SpoT. J Bacteriol. 2007 Sep;189(17):6140-7. Epub 2007 Jul 6. PMID:17616600 doi:http://dx.doi.org/10.1128/JB.00315-07
  35. ↑ Foti JJ, Persky NS, Ferullo DJ, Lovett ST. Chromosome segregation control by Escherichia coli ObgE GTPase. Mol Microbiol. 2007 Jul;65(2):569-81. Epub 2007 Jun 18. PMID:17578452 doi:http://dx.doi.org/10.1111/j.1365-2958.2007.05811.x
  36. ↑ Persky NS, Ferullo DJ, Cooper DL, Moore HR, Lovett ST. The ObgE/CgtA GTPase influences the stringent response to amino acid starvation in Escherichia coli. Mol Microbiol. 2009 Jul;73(2):253-66. doi: 10.1111/j.1365-2958.2009.06767.x. Epub, 2009 Jun 23. PMID:19555460 doi:http://dx.doi.org/10.1111/j.1365-2958.2009.06767.x
  37. ↑ Gkekas S, Singh RK, Shkumatov AV, Messens J, Fauvart M, Verstraeten N, Michiels J, Versees W. Structural and Biochemical Analysis of Escherichia coli ObgE, a Central Regulator of Bacterial Persistence. J Biol Chem. 2017 Feb 21. pii: jbc.M116.761809. doi: 10.1074/jbc.M116.761809. PMID:28223358 doi:http://dx.doi.org/10.1074/jbc.M116.761809

5m04, resolution 1.85Å

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