Molecular Bacteriology

Prof R K Poole - West Riding Chair in Microbiology

Microbial respiration.

We are trying to obtain a comprehensive understanding of how bacteria respire - aerobically and anaerobically - and how the diverse oxygen-reactive proteins of bacteria are involved in respiratory electron transfer, protection of bacteria from oxidative stress and the sensing of environmental and cellular oxygen levels.

Problems of special interest at present are

a Systems Biology approach to understanding oxygen metabolism in E. coli (an international consortium in the SysMO2 Initiative involving Sheffield, Amsterdam, Magdeburg and Stuttgart),
haemoglobins in various bacteria, especially pathogens, that bind oxygen but also allow detoxification of nitric oxide (NO, see below) and
the antibacterial activity of the respiratory poison carbon monoxide (CO) and molecules that release CO in physiologically tractable ways (CORMs).

This last project is a collaborative venture with Dr Roberto Motterlini (Italian Institute of Technology) and Professor Brian Mann (Chemistry, Sheffield) and seeks to understand the balance between CO (derived from the host or by clinical intervention), pathogenic bacteria and the inflammatory response.

Microbial resistance to oxidative and nitrosative stresses.

Of particular interest at present are those mechanisms that allow bacteria to survive the presence of NO - a simple diatomic radical, but which is immensely important throughout biology. NO is not only a physiologically critical signalling molecule involved in cardiovascular function, but also a potent product of the antibacterial activity involved in innate immunity.

We have made major advances in understanding the function, regulation and properties of the flavohaemoglobin (Hmp) of Escherichia coli and Salmonella: the only confirmed function of this protein is to enzymically detoxify NO. We are also studying respiratory and NO metabolism in bacteria of clinical and pathogenic significance (especially Campylobacter, Mycobacterium and Salmonella) and this work is often done collaboratively with Professor Robert Read (Royal Hallamshire Hospital).

Figure Legend - The flavohaemoglobin (HMP) of Escherichia coli is an oxygen-binding protein whose synthesis is strongly up-regulated by nitric oxide (NO) and oxidative stress. Its possible roles in oxygen or NO sensing are being studied using molecular genetic, physiological and protein chemical methods.

Selected Publications

SHEPHERD, M., BERNHARDT, P. V. & POOLE, R. K. Globin-mediated nitric oxide detoxification in the foodborne pathogenic bacterium Campylobacter jejuni proceeds via a dioxygenase or denitrosylase mechanism. Nitric Oxide: Biol. Chem., in the press

McLEAN, S., BOWMAN, L. A. H. & POOLE, R. K. (2010) Peroxynitrite stress is exacerbated by flavohaemoglobin-derived oxidative stress in Salmonella typhimurium and is relieved by nitric oxide. Microbiology 156, 3556-3565

WILSON, J. L., JESSE, H. E., POOLE, R. K. & DAVIDGE, K. S. (2010) Antibacterial effects of carbon monoxide. Curr. Pharmaceut. Biotechnol., in the press

SHEPHERD, M. & POOLE, R. K. (2010) Bacterial respiratory chains. In Encyclopedia of Biophysics (Ed. G. C. K. Roberts), Springer-European Biophysical Societies Association (EBSA). In press

POOLE, R. K. & SHEPHERD, M. (2010) Bacterial globins. In Encyclopedia of Biophysics (Ed. G. C. K. Roberts), Springer-European Biophysical Societies Association (EBSA). In press

THOMAS, M., SHEPHERD, M., POOLE, R. K., VAN VLIET, A. H. M., KELLY, D. J., & PEARSON, B. M. (2010) Two respiratory enzyme systems in Campylobacter jejuni NCTC 11168 contribute to growth on L-lactate. Environ. Microbiol. doi: 10.1111/j.1462-2920.2010.02307.x

McLEAN, S., BOWMAN, L. A. H., SANGUINETTI, G., READ, R. C. & POOLE, R. K. (2010) Peroxynitrite toxicity in Escherichia coli K12 elicits expresion of oxidative stress responses and protein nitration and nitrosylation. J. Biol. Chem. 285, 20724-20731

SHEPHERD M., SANGUINETTI, G., COOK, G. M. & POOLE, R. K. (2010) Compensations for diminished terminal oxidase activity in Escherichia coli: cytochrome bd-II-mediated respiration and glutamate metabolism. J. Biol. Chem. 285, 18464-18472

SVENSSON, L., POLJAKOVIC, M., SÄVE, S., GILBERTHORPE, N., SCHÖN, T., STRID, S., CORKER, H., POOLE, R. K. & PERSSON, K. (2010) Role of flavohemoglobin in combating nitrosative stress in uropathogenic Escherichia coli – implications for urinary tract infection. Microb. Pathogen. 49, 59-66

McLEAN, S., BOWMAN, L. A. H. & POOLE, R. K. (2010) KatG from Salmonella typhimurium is a peroxynitritase. FEBS Lett. 584, 1628-1632

SHEPHERD M., BARYNIN, V., LU, C., BERNHARDT, P. V., WU, G., YEH, S.-R., EGAWA, T., SEDELNIKOVA, S. E., RICE, D. W., WILSON, J. L. & POOLE , R. K. (2010) The single-domain globin from the pathogenic bacterium Campylobacter jejuni: resolution of the D-helix, influence of proximal hydrogen bonding on ligand binding, and peroxidase-like redox properties. J. Biol. Chem. 285, 12747-12754

DAVIDGE, K. S., MOTTERLINI, R., MANN, B. E., WILSON J. L. & POOLE, R. K. (2009) Carbon monoxide in biology and microbiology: surprising roles for the "Detroit Perfume". Adv. Microb. Physiol. 56, 85-167

LATIMER, J., STOKES, S. L., GRAHAM, A. I., BUNCH, J., JACKSON, R. J., McLEOD, C. W. & POOLE, R. K. (2009) A novel method for exploring elemental composition of microbial communities: laser ablation-inductively coupled plasma-mass spectrometry of intact bacterial colonies. J. Microbiol. Meth. 79, 329-335.

LAVER, J. R. STEVANIN, T. M., MESSENGER, S. L., LUNN, A. D., LEE, M. E., MOIR, J. W. B., POOLE, R. K. & READ, R. C. (2009) Bacterial nitric oxide detoxification prevents host cell S-nitrosothiol formation: a novel mechanism of bacterial pathogenesis. FASEB J. 24, 286-295.

GRAHAM, A. I. HUNT, S., STOKES, S. L., BRAMALL, N., BUNCH, J., COX, A. G., MCLEOD, C. W. & POOLE, R. K. (2009) Severe zinc depletion of Escherichia coli: roles for high-affinity zinc binding by ZinT, zinc transport and zinc-independent proteins. J. Biol. Chem. 284, 18377-18389.

MALEKI-DIZAJI, S., HOLCOMBE, M., ROLFE, M. D., FISHER, P., GREEN, J. POOLE, R. K. & GRAHAM, A. I. (2009) A systematic approach to understanding Escherichia coli responses to oxygen: from microarray raw data to pathways and published abstracts. Online J. Bioinformatics 10, 51-59.

DAVIDGE, K. S. SANGUINETTI, G., YEE, C. H. COX, A. G., MCLEOD, C. W., MONK, C. E., MANN, B. E., MOTTERLINI, R. & AND POOLE, R. K. (2009) Carbon monoxide-releasing antibacterial molecules target respiration and global transcriptional regulators. J. Biol. Chem. 284, 4516-4524.

DESMARD, M., DAVIDGE, K. S., BOUVET, O., MORIN, D., ROUX, D., RICARD, J. D., DENAMUR, E., POOLE, R. K., MONTRAVERS, P., MOTTERLINI, R. & BOCZOWSKI, J. (2009) A carbon monoxide-releasing molecule (CORM-3) exerts bactericidal activity against Pseudomonas aeruginosa and improves survival in an animal model of bacteraemia. FASEB J, 23, 1023-1031.

MASON, M. G., SHEPHERD, M., NICHOLLS, P., DOBBIN, P. S., DODSWORTH, K. S. POOLE, R. K. & COOPER, C. E. (2008) Cytochrome bd confers nitric oxide resistance to E. coli. Nature Chem. Biol. 5, 94-96.

JACKSON, R. J., BINET, M. R. B., LEE, L. J., MA, R., GRAHAM, A. I., McLEOD, C. W. & POOLE, R. K. (2008) Expression of the PitA phosphate/metal transporter of Escherichia coli is responsive to zinc and inorganic phosphate levels. FEMS Microbiol. Lett. 289, 219-224.

MONK, C. E., PEARSON, B. M., MULHOLLAND, F. & POOLE, R. K. (2008) Oxygen- and NssR-dependent globin expression and enhanced iron acquisition in the response of Campylobacter to nitrosative stress. J. Biol. Chem., 283, 28413-28425.

POOLE, R. K. (2008) Microbial haemoglobins – proteins at the crossroads of oxygen and nitric oxide metabolism. In: Dioxygen Binding and Sensing Proteins. A Tribute to Beatrice and Jonathan Wittenberg. (Bolognesi, M., Di Prisco, G. & Verde, C.). Springer-Verlag Italia

GILBERTHORPE, N. J. & POOLE, R. K. (2008) Nitric oxide homeostasis in Salmonella typhimurium: roles of respiratory nitrate reductase and flavohemoglobin. J. Biol. Chem. 283, 11146-11154.

GRUIA, F., KUBO, M., YE, X., IONASCU, D., LU, C., POOLE, R. K., YEH, S.-R. & CHAMPION, P. (2008) Coherence spectroscopy investigations of the low-frequency vibrations of heme: Effects of protein-specific perturbations. J. Am. Chem. Soc. 130, 5231-5244.

PARTRIDGE, J. D., BROWNING, D. F., XU, M., NEWNHAM, L. J., SCOTT, C., ROBERTS, R. E., POOLE, R, K. & GREEN, J. (2008) Characterization of the Escherichia coli K-12 ydhYVWXUT operon: regulation by FNR, NarL and NarP. Microbiology 154, 608-618.

LU, C., EGAWA, T., MUKAI, M., POOLE, R. K. & YEH, S.-R. (2008) Hemoglobins from Mycobacterium tuberculosis and Campylobacter jejuni: A comparative study with resonance Raman spectroscopy. In: Methods in Enzymology. vol. 437. Globins and Other NO-Reactive Proteins, Part B, pp. 255-286. San Diego: Elsevier.

PULLAN, S. T., MONK, C. E., LEE, L. & POOLE, R. K. (2008) Microbial responses to nitric oxide and nitrosative stress: growth, 'omic' and physiological methods. In: Methods in Enzymology. vol. 437. Globins and Other NO-Reactive Proteins, Part B, pp. 499-519. San Diego: Elsevier.

PICKFORD, J. L., WAINWRIGHT, L, WU, G. & POOLE, R. K. (2008) Expression and purification of Cgb and Ctb, the NO-inducible globins of the foodborne bacterial pathogen Campylobacter jejuni. In: Methods in Enzymology. vol. 436. Globins and Other NO-Reactive Proteins, Part A, pp. 289-302. San Diego: Elsevier.

LEWIS, M. E. S, CORKER, H. A., GOLLAN, B. & POOLE, R. K. (2008) A survey of methods for the purification of microbial flavohemoglobins. In: Methods in Enzymology. vol. 436. Globins and Other NO-Reactive Proteins, Part A, pp. 169-186. San Diego: Elsevier.

KALNENIEKS, U., GALININA, N., STRAZDINA, I., KRAVALE, Z., PICKFORD, J. L. RUTKIS, R. & POOLE, R. K. (2008) NADH dehydrogenase deficiency results in low respiration rates and improved aerobic growth of Zymomonas mobilis. Microbiology. 154, 989-994.