IGEM:Caltech/2008/Project/ROS

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iGEM 2008

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Conjugation

  • Can get conjugation from Salmonella to E. coli [1]
    • Papers seem to assume that conjugation is quick, and maintenance is the hard part (unless we engineer a shuttle vector). Doesn't address specificity, though.
  • "Conjugation of S. typhi with E. coli F+ carrying P1CM+ gave three types of S. typhi CMr clones: those which carry the whole P1CMphage, those with the PldCM element, and those with nontransferable CMr."[2]
  • Can get transfer of an F' plasmid from E. coli to Salmonella[3]
  • Doug Tischer 21:12, 26 May 2008 (EDT)There exists a plasmid (pAT187) that can be transfered from E. coli to a wide range of gram-positive bacteria [4]
  • Doug Tischer 17:42, 27 May 2008 (EDT)Surface exclusion seems to be the name given to the process that prevents a bacteria containing a plasmid from conjugating and receiving the same plasmid again. In F plasmid, the are the traS and traT genes. traT inhibits the formation of mating aggregates. [5]
  • Doug Tischer 17:51, 27 May 2008 (EDT) Biobrick part BBa_J01000 (TraJf) is a transcriptional activator that turns on the expression of sex pili forming genes off of the F plasmid. This could be used to trigger the oxidative burst upon conjugation instead. See this page for information explaining this and other conjugation related Biobricks.

Quorum Sensing

  • Given the complexity of a conjugation based triggering system, perhaps quorum sensing would be more feasible. Interspecies signaling is carried out through acyl-homoserine lactone (AHL). These are very specific, and each type of bacteria produces there own unique one. It only takes one cytoplasmic receptor protein to bind to the AHL and activate transcription. The production of AHL is associated with a switch from a benign to a pathogenic form in several bacteria. We could make our cells target Salmonella by giving them the receptor for the Salmonella AHL. The only thing we would need to change is to make the detection quite sensitive, so that our cells kill the Salmonella become pathogenic. [6]
  • lsrB [7]

Specificity

  • Could we detect transmission of the Salmonella virulence plasmid?
    • "The second experiment was to physically observe the transferred virulence plasmid in the recipient strain.... Therefore, E. coli was chosen to be the recipient in matings with Salmonella.... Transconjugants were obtained at a frequency of approximately 2 × 10−5 transconjugants/donor.... Ethidium bromide staining of the pulsed-field gel shows that the restriction pattern of the transconjugant genomes matches that of the E. coli recipient, except for the acquisition of the virulence plasmid from the Salmonella donor. The identity of the virulence plasmid was confirmed by subsequent Southern hybridization with the kanamycin resistance gene of MudJ.... This is clear evidence that conjugation has occurred."[8]
    • sdiA transcription factor? [9]
    • Or spvR[10, 11]. It functions in E. coli and is necessary for virulence. So our bacteria are floating around waiting for conjugation from a pathogenic Salmonella. Once that happens, a spvR promoter turns on, triggering the ROS. The low conjugation frequency is troubling. However, some of that might be low efficiency genomic integration.

H2O2 Transport

  • Diffusion of H2O2 across E. coli membranes is "exhibit substantial, but limited, permeability to H2O2." The review suggested some aquaporins in yeast increase cell membrane permeability to H2O2. [12]
  • The strain JI377 (a deletion for katE, katG, and ahp) cannot scavange H2O2 whatsoever and excretes it into the medium. [13]

References

  1. Makanera A, Arlet G, Gautier V, and Manai M. Molecular epidemiology and characterization of plasmid-encoded beta-lactamases produced by Tunisian clinical isolates of Salmonella enterica serotype Mbandaka resistant to broad-spectrum cephalosporins. J Clin Microbiol. 2003 Jul;41(7):2940-5. PubMed ID:12843024 | HubMed [makanera]
  2. Kondo E and Mitsuhashi S. Drug resistance of enteric bacteria. VI. Introduction of bacteriophage P1CM into Salmonella typhi and formation of PldCM and F-CM elements. J Bacteriol. 1966 May;91(5):1787-94. PubMed ID:5327907 | HubMed [kondo]
  3. Lenny AB and Margolin P. Locations of the opp and supX genes of Salmonella typhimurium and Escherichia coli. J Bacteriol. 1980 Aug;143(2):747-52. PubMed ID:7009564 | HubMed [lenny]
  4. Mazodier P, Petter R, and Thompson C. Intergeneric conjugation between Escherichia coli and Streptomyces species. J Bacteriol. 1989 Jun;171(6):3583-5. PubMed ID:2656662 | HubMed [Mazodier]
  5. Achtman M, Morelli G, and Schwuchow S. Cell-cell interactions in conjugating Escherichia coli: role of F pili and fate of mating aggregates. J Bacteriol. 1978 Sep;135(3):1053-61. PubMed ID:357413 | HubMed [Achtman]
  6. Federle MJ and Bassler BL. Interspecies communication in bacteria. J Clin Invest. 2003 Nov;112(9):1291-9. DOI:10.1172/JCI20195 | PubMed ID:14597753 | HubMed [Federle]
  7. Miller ST, Xavier KB, Campagna SR, Taga ME, Semmelhack MF, Bassler BL, and Hughson FM. Salmonella typhimurium recognizes a chemically distinct form of the bacterial quorum-sensing signal AI-2. Mol Cell. 2004 Sep 10;15(5):677-87. DOI:10.1016/j.molcel.2004.07.020 | PubMed ID:15350213 | HubMed [miller]
  8. Ahmer BM, Tran M, and Heffron F. The virulence plasmid of Salmonella typhimurium is self-transmissible. J Bacteriol. 1999 Feb;181(4):1364-8. PubMed ID:9973370 | HubMed [ahmer]
  9. Ahmer BM, van Reeuwijk J, Timmers CD, Valentine PJ, and Heffron F. Salmonella typhimurium encodes an SdiA homolog, a putative quorum sensor of the LuxR family, that regulates genes on the virulence plasmid. J Bacteriol. 1998 Mar;180(5):1185-93. PubMed ID:9495757 | HubMed [ahmer2]
  10. Grob P, Kahn D, and Guiney DG. Mutational characterization of promoter regions recognized by the Salmonella dublin virulence plasmid regulatory protein SpvR. J Bacteriol. 1997 Sep;179(17):5398-406. PubMed ID:9286993 | HubMed [grob]
  11. Wilson JA, Doyle TJ, and Gulig PA. Exponential-phase expression of spvA of the Salmonella typhimurium virulence plasmid: induction in intracellular salts medium and intracellularly in mice and cultured mammalian cells. Microbiology. 1997 Dec;143 ( Pt 12):3827-39. DOI:10.1099/00221287-143-12-3827 | PubMed ID:9421907 | HubMed [wilson]
  12. Bienert GP, Schjoerring JK, and Jahn TP. Membrane transport of hydrogen peroxide. Biochim Biophys Acta. 2006 Aug;1758(8):994-1003. DOI:10.1016/j.bbamem.2006.02.015 | PubMed ID:16566894 | HubMed [Bienert]
  13. Seaver LC and Imlay JA. Alkyl hydroperoxide reductase is the primary scavenger of endogenous hydrogen peroxide in Escherichia coli. J Bacteriol. 2001 Dec;183(24):7173-81. DOI:10.1128/JB.183.24.7173-7181.2001 | PubMed ID:11717276 | HubMed [Seaver]
  14. Plainkum P, Fuchs SM, Wiyakrutta S, and Raines RT. Creation of a zymogen. Nat Struct Biol. 2003 Feb;10(2):115-9. DOI:10.1038/nsb884 | PubMed ID:12496934 | HubMed [Plainkum]
  15. Firbank SJ, Rogers MS, Wilmot CM, Dooley DM, Halcrow MA, Knowles PF, McPherson MJ, and Phillips SE. Crystal structure of the precursor of galactose oxidase: an unusual self-processing enzyme. Proc Natl Acad Sci U S A. 2001 Nov 6;98(23):12932-7. DOI:10.1073/pnas.231463798 | PubMed ID:11698678 | HubMed [Firbank]
  16. Sun L, Petrounia IP, Yagasaki M, Bandara G, and Arnold FH. Expression and stabilization of galactose oxidase in Escherichia coli by directed evolution. Protein Eng. 2001 Sep;14(9):699-704. PubMed ID:11707617 | HubMed [Sun]
  17. Uehara Y, Kikuchi K, Nakamura T, Nakama H, Agematsu K, Kawakami Y, Maruchi N, and Totsuka K. H(2)O(2) produced by viridans group streptococci may contribute to inhibition of methicillin-resistant Staphylococcus aureus colonization of oral cavities in newborns. Clin Infect Dis. 2001 May 15;32(10):1408-13. DOI:10.1086/320179 | PubMed ID:11317240 | HubMed [Uehara]
  18. Toomey D and Mayhew SG. Purification and characterisation of NADH oxidase from Thermus aquaticus YT-1 and evidence that it functions in a peroxide-reduction system. Eur J Biochem. 1998 Feb 1;251(3):935-45. PubMed ID:9490070 | HubMed [Toomey]
  19. Whittaker JW. The radical chemistry of galactose oxidase. Arch Biochem Biophys. 2005 Jan 1;433(1):227-39. DOI:10.1016/j.abb.2004.08.034 | PubMed ID:15581579 | HubMed [Whittaker]
All Medline abstracts: PubMed | HubMed