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Crystal structures of Mycobacterial MeaB and MMAA-like GTPases

  • Published:
Journal of Structural and Functional Genomics

Abstract

The methylmalonyl Co-A mutase-associated GTPase MeaB from Methylobacterium extorquens is involved in glyoxylate regulation and required for growth. In humans, mutations in the homolog methylmalonic aciduria associated protein (MMAA) cause methylmalonic aciduria, which is often fatal. The central role of MeaB from bacteria to humans suggests that MeaB is also important in other, pathogenic bacteria such as Mycobacterium tuberculosis. However, the identity of the mycobacterial MeaB homolog is presently unclear. Here, we identify the M. tuberculosis protein Rv1496 and its homologs in M. smegmatis and M. thermoresistibile as MeaB. The crystal structures of all three homologs are highly similar to MeaB and MMAA structures and reveal a characteristic three-domain homodimer with GDP bound in the G domain active site. A structure of Rv1496 obtained from a crystal grown in the presence of GTP exhibited electron density for GDP, suggesting GTPase activity. These structures identify the mycobacterial MeaB and provide a structural framework for therapeutic targeting of M. tuberculosis MeaB.

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Abbreviations

ALS:

Advanced Light Source

APF:

Atomic property force

ASU:

Asymmetric unit

GTP:

Guanosine 5′ triphosphate

IMAC:

Immobilized metal ion affinity chromatography

MCM:

Methylmalonyl-CoA-mutase

MMAA:

Methylmalonyl aciduria associated protein A

PDB:

Protein Data Bank

SBM:

Sleeping beauty mutase

SSGCID:

Seattle Structural Genomics Center for Infectious Disease

References

  1. Korotkova N, Lidstrom ME (2004) MeaB is a component of the methylmalonyl-CoA mutase complex required for protection of the enzyme from inactivation. J Biol Chem 279:13652–13658

    Article  CAS  PubMed  Google Scholar 

  2. Korotkova N, Chistoserdova L, Kuksa V, Lidstrom ME (2002) Glyoxylate regeneration pathway in the methylotroph Methylobacterium extorquens AM1. J Bacteriol 184:1750–1758

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  3. Padovani D, Banerjee R (2009) A G-protein editor gates coenzyme B12 loading and is corrupted in methylmalonic aciduria. Proc Natl Acad Sci USA 106:21567–21572

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  4. Dobson CM, Wai T, Leclerc D, Kadir H, Narang M, Lerner-Ellis JP, Hudson TJ, Rosenblatt DS, Gravel RA (2002) Identification of the gene responsible for the cblB complementation group of vitamin B12-dependent methylmalonic aciduria. Hum Mol Genet 11:3361–3369

    Article  CAS  PubMed  Google Scholar 

  5. Hubbard PA, Padovani D, Labunska T, Mahlstedt SA, Banerjee R, Drennan CL (2007) Crystal structure and mutagenesis of the metallochaperone MeaB: insight into the causes of methylmalonic aciduria. J Biol Chem 282:31308–31316

    Article  CAS  PubMed  Google Scholar 

  6. Froese DS, Kochan G, Muniz JR, Wu X, Gileadi C, Ugochukwu E, Krysztofinska E, Gravel RA, Oppermann U, Yue WW (2010) Structures of the human GTPase MMAA and vitamin B12-dependent methylmalonyl-CoA mutase and insight into their complex formation. J Biol Chem 285:38204–38213

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  7. Savvi S, Warner DF, Kana BD, McKinney JD, Mizrahi V, Dawes SS (2008) Functional characterization of a vitamin B12-dependent methylmalonyl pathway in Mycobacterium tuberculosis: implications for propionate metabolism during growth on fatty acids. J Bacteriol 190:3886–3895

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  8. Celis RT, Leadlay PF, Roy I, Hansen A (1998) Phosphorylation of the periplasmic binding protein in two transport systems for arginine incorporation in Escherichia coli K-12 is unrelated to the function of the transport system. J Bacteriol 180:4828–4833

    PubMed Central  CAS  PubMed  Google Scholar 

  9. Froese DS, Dobson CM, White AP, Wu X, Padovani D, Banerjee R, Haller T, Gerlt JA, Surette MG, Gravel RA (2009) Sleeping beauty mutase (sbm) is expressed and interacts with ygfd in Escherichia coli. Microbiol Res 164:1–8

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  10. Aslanidis C, de Jong PJ (1990) Ligation-independent cloning of PCR products (LIC-PCR). Nucleic Acids Res 18:6069–6074

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  11. Studier FW (2005) Protein production by auto-induction in high density shaking cultures. Protein Expr Purif 41:207–234

    Article  CAS  PubMed  Google Scholar 

  12. Alexandrov A, Dutta K, Pascal SM (2001) MBP fusion protein with a viral protease cleavage site: one-step cleavage/purification of insoluble proteins. Biotechniques 30:1194–1198

    CAS  PubMed  Google Scholar 

  13. Kabsch W (2010) Xds. Acta Crystallogr D Biol Crystallogr 66:125–132

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  14. Vagin AA, Teplyakov A (1997) MOLREP: an automated program for molecular replacement. J Appl Cryst 30:1022–1025

    Article  CAS  Google Scholar 

  15. Collaborative Computational Project, Number 4 (1994) The CCP4 suite: programs for protein crystallography. Acta Crystallogr D Biol Crystallogr 50:760–763

  16. Murshudov GN, Vagin AA, Dodson EJ (1997) Refinement of macromolecular structures by the maximum-likelihood method. Acta Crystallogr D Biol Crystallogr 53:240–255

    Article  CAS  PubMed  Google Scholar 

  17. Emsley P, Cowtan K (2004) Coot: model-building tools for molecular graphics. Acta Crystallogr D Biol Crystallogr 60:2126–2132

    Article  PubMed  Google Scholar 

  18. McCoy AJ, Grosse-Kunstleve RW, Adams PD, Winn MD, Storoni LC, Read RJ (2007) Phaser crystallographic software. J Appl Crystallogr 40:658–674

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  19. Chen VB, Arendall WB 3rd, Headd JJ, Keedy DA, Immormino RM, Kapral GJ, Murray LW, Richardson JS, Richardson DC (2010) MolProbity: all-atom structure validation for macromolecular crystallography. Acta Crystallogr D Biol Crystallogr 66:12–21

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  20. Myler PJ, Stacy R, Stewart L, Staker BL, Van Voorhis WC, Varani G, Buchko GW (2009) The Seattle Structural Genomics Center for Infectious Disease (SSGCID). Infect Disord Drug Targets 9:493–506

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  21. Stacy R, Begley DW, Phan I, Staker BL, Van Voorhis WC, Varani G, Buchko GW, Stewart LJ, Myler PJ (2011) Structural genomics of infectious disease drug targets: the SSGCID. Acta Crystallogr Sect F Struct Biol Cryst Commun 67:979–984

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  22. Baugh L, Gallagher LA, Patrapuvich R, Clifton MC, Gardberg AS, Edwards TE, Armour B, Begley DW, Dieterich SH, Dranow DM, Abendroth J, Fairman JW, Fox D 3rd, Staker BL, Phan I, Gillespie A, Choi R, Nakazawa-Hewitt S, Nguyen MT, Napuli A, Barrett L, Buchko GW, Stacy R, Myler PJ, Stewart LJ, Manoil C, Van Voorhis WC (2013) Combining functional and structural genomics to sample the essential Burkholderia structome. PLoS One 8:e53851

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  23. Altaf M, Miller CH, Bellows DS, O’Toole R (2010) Evaluation of the Mycobacterium smegmatis and BCG models for the discovery of Mycobacterium tuberculosis inhibitors. Tuberculosis (Edinb) 90:333–337

    Article  CAS  Google Scholar 

  24. Edwards TE, Liao R, Phan I, Myler PJ, Grundner C (2012) Mycobacterium thermoresistibile as a source of thermostable orthologs of Mycobacterium tuberculosis proteins. Protein Sci 21:1093–1096

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  25. Totrov M (2011) Ligand binding site superposition and comparison based on atomic property fields: identification of distant homologues, convergent evolution and PDB-wide clustering of binding sites. BMC Bioinform 12(Suppl 1):S35

    Article  Google Scholar 

  26. Totrov M (2008) Atomic property fields: generalized 3D pharmacophoric potential for automated ligand superposition, pharmacophore elucidation and 3D QSAR. Chem Biol Drug Des 71:15–27

    Article  CAS  PubMed  Google Scholar 

  27. Giganti D, Guillemain H, Spadoni JL, Nilges M, Zagury JF, Montes M (2010) Comparative evaluation of 3D virtual ligand screening methods: impact of the molecular alignment on enrichment. J Chem Inf Model 50:992–1004

    Article  CAS  PubMed  Google Scholar 

  28. Lofgren M, Padovani D, Koutmos M, Banerjee R (2013) A switch III motif relays signaling between a B12 enzyme and its G-protein chaperone. Nat Chem Biol 9:535–539

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  29. Lofgren M, Koutmos M, Banerjee R (2013) Autoinhibition and signaling by the switch II motif in the G-protein chaperone of a radical B12 enzyme. J Biol Chem 288:30980–30989

    Article  PubMed Central  CAS  PubMed  Google Scholar 

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Acknowledgments

The authors thank the whole SSGCID team. This research was funded with federal funds from the National Institute of Allergy and Infectious Diseases National Institutes of Health, Department of Health and Human Services under Contract Nos. HHSN272201200025C and HHSN272200700057C. All data sets except for the Mtb Rv1496-GDP (3MD0) complex were obtained through the ALS Collaborative Crystallography program. The Berkeley Center for Structural Biology is supported in part by the National Institutes of Health, National Institute of General Medical Sciences, and the Howard Hughes Medical Institute. The Advanced Light Source is supported by the Director, Office of Science, Office of Basic Energy Sciences, and the U.S. Department of Energy under Contract No. DE-AC02-05CH11231.

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Authors and Affiliations

  1. Beryllium, Seattle Structural Genomics Center for Infectious Disease (SSGCID), Bainbridge Island, WA, 98110, USA

    Thomas E. Edwards, Jameson Bullen, Ruth O. Baydo, Pam Witte, Jan Abendroth, Matthew C. Clifton & Donald D. Lorimer

  2. Beryllium, Seattle Structural Genomics Center for Infectious Disease (SSGCID), Bedford, MA, 01730, USA

    Thomas E. Edwards, Jameson Bullen, Ruth O. Baydo, Pam Witte, Jan Abendroth, Matthew C. Clifton & Donald D. Lorimer

  3. Seattle Biomedical Research Institute, Seattle Structural Genomics Center for Infectious Disease (SSGCID), Seattle, WA, 98109, USA

    Loren Baugh, Kaitlin Thompkins, Isabelle Q. H. Phan, Peter J. Myler, Bart L. Staker & Christoph Grundner

  4. Berkeley Center for Structural Biology, Ernest Orlando Lawrence Berkeley National Laboratory, Berkeley, CA, 94720, USA

    Banumathi Sankaran

  5. Departments of Medicine, Microbiology, and Global Health, University of Washington, Seattle Structural Genomics Center for Infectious Disease (SSGCID), Seattle, WA, 98195, USA

    Wesley C. Van Voorhis

  6. Department of Medical Education and Biomedical Information, University of Washington, Seattle, WA, 98195, USA

    Peter J. Myler

  7. Department of Global Health, University of Washington, Seattle, WA, 98195, USA

    Peter J. Myler

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  1. Thomas E. Edwards
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  2. Loren Baugh
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  3. Jameson Bullen
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  5. Pam Witte
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  7. Isabelle Q. H. Phan
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  8. Jan Abendroth
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  9. Matthew C. Clifton
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  10. Banumathi Sankaran
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  11. Wesley C. Van Voorhis
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  12. Peter J. Myler
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  13. Bart L. Staker
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  14. Christoph Grundner
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  15. Donald D. Lorimer
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Correspondence to Thomas E. Edwards.

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Edwards, T.E., Baugh, L., Bullen, J. et al. Crystal structures of Mycobacterial MeaB and MMAA-like GTPases. J Struct Funct Genomics 16, 91–99 (2015). https://doi.org/10.1007/s10969-015-9197-2

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  • DOI: https://doi.org/10.1007/s10969-015-9197-2

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