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Kenan C Murphy PhD

TitleAssistant Professor
InstitutionUniversity of Massachusetts Medical School
DepartmentMicrobiology and Physiological Systems
AddressUniversity of Massachusetts Medical School
55 Lake Avenue North
Worcester MA 01655
Phone508-856-6042
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    Other Positions
    InstitutionUMMS - School of Medicine
    DepartmentMicrobiology and Physiological Systems

    InstitutionUMMS - Graduate School of Biomedical Sciences
    DepartmentMolecular Genetics and Microbiology

    InstitutionUMMS - Programs, Centers and Institutes
    DepartmentBacterial Genetics and Pathogenesis


    Collapse Biography 
    Collapse education and training
    Catholic University of America, Washington, DC, United StatesBAPhysics
    University of Maryland, College Park, College Park, MD, United StatesPHDBiochemistry

    Collapse Overview 
    Collapse overview


    Academic Background



    Ph. D. (1983) University of Maryland



    Recombineering technology for gene replacement in bacterial pathogens



    Identification of Drug Targets in M. tuberculosis - creating regulatable strains for use in whole cell screens with small molecules



    My work invovles the use of Red recombineering technology for gene replacement in bacterial pathogens. My lab was the first to show that the lambda Red recombination system promotes gene replacement of electroporated linear DNA substrates into the Escherichia coliK-12 chromosome at a very high efficiency (Murphy, 1998). The system is also useful in pathogenic species of E. coli (Murphy & Campellone, 2003). Work continues to improve recombinering technology in pathogens such as Pseudomonas aeruginosa and Mycobacterium tuberculosis by expression of Red-lke recombination systems from phage known to infect these hosts.



     



    My lab is also interested in the mechnaism of the bacteriophage lambda Red recombination system. The system consist of two proteins, the ssDNA annealing Bet protein and the 5’-3’ dsDNA lambda exonuclease. These two proteins form a complex in vitro, and are thought to interact with each other in vivo. We have isolated various mutants of Bet that are deficient for both recombination and recombineering, and some that are deficient for one but not the other.   



     



     


    Proposed mechanism of Red Recombineering



    Lambda Exo's 5' exonuclease activity (red trapezoid)) generates ssDNA, which serves as a substrate for lambda Bet (blue oligomeric ring) to bind and promote annealing to ssDNA in the lagging strand of a replication fork. This structure is stabilized by the lambda Beta protein, unitl another fork comes by and generates both a wild type and a recombinant chromosome.



    replication Kenan Murphy figure page



    Collapse Rotation Projects


    Rotation Projects





    1. Study the interaction of the ssDNA annealing protein Bet and its cognate exonuclease with dsDNA ends. Compare wild type Bet with one or mutant Bet proteins.  Use both genetic and biochemical studies to answer the question of how this annealase binds to ssDNA and promotes recombination.

       




    2. Develop recombineering technologies for M. tuberculosis involving the electroporation of oligos containing clinically relevant single nucleotide polymorphisms (SNPs).

       




    3. Generate a recombineering system for Pseudomonas aeruginosa via the electroporation of PCR substrates into electrocompetent cells containing PA phage Red-like recombination systems. 




     



    Collapse Bibliographic 
    Collapse selected publications
    Publications listed below are automatically derived from MEDLINE/PubMed and other sources, which might result in incorrect or missing publications. Faculty can login to make corrections and additions.
    List All   |   Timeline
    1. Prigozhin DM, Papavinasasundaram KG, Baer CE, Murphy KC, Moskaleva A, Chen TY, Alber T, Sassetti CM. Structural and Genetic Analyses of the Mycobacterium tuberculosis Protein Kinase B Sensor Domain Identify a Potential Ligand-binding Site. J Biol Chem. 2016 Oct 28; 291(44):22961-22969. PMID: 27601474.
      View in: PubMed
    2. Warrier T, Kapilashrami K, Argyrou A, Ioerger TR, Little D, Murphy KC, Nandakumar M, Park S, Gold B, Mi J, Zhang T, Meiler E, Rees M, Somersan-Karakaya S, Porras-De Francisco E, Martinez-Hoyos M, Burns-Huang K, Roberts J, Ling Y, Rhee KY, Mendoza-Losana A, Luo M, Nathan CF. N-methylation of a bactericidal compound as a resistance mechanism in Mycobacterium tuberculosis. Proc Natl Acad Sci U S A. 2016 Aug 02; 113(31):E4523-30. PMID: 27432954.
      View in: PubMed
    3. Murphy KC. ? Recombination and Recombineering. EcoSal Plus. 2016 May; 7(1). PMID: 27223821.
      View in: PubMed
    4. Murphy, KC. EcoSal Plus 2016; doi:10.1128/ecosalplus.ESP-0011-2015. Phage Lambda Recombination and Recombineering. 2016.
    5. Nambi S, Long JE, Mishra BB, Baker R, Murphy KC, Olive AJ, Nguyen HP, Shaffer SA, Sassetti CM. The Oxidative Stress Network of Mycobacterium tuberculosis Reveals Coordination between Radical Detoxification Systems. Cell Host Microbe. 2015 Jun 10; 17(6):829-37. PMID: 26067605.
      View in: PubMed
    6. Murphy KC, Papavinasasundaram K, Sassetti CM. Mycobacterial recombineering. Methods Mol Biol. 2015; 1285:177-99. PMID: 25779316.
      View in: PubMed
    7. Meniche X, Otten R, Siegrist MS, Baer CE, Murphy KC, Bertozzi CR, Sassetti CM. Subpolar addition of new cell wall is directed by DivIVA in mycobacteria. Proc Natl Acad Sci U S A. 2014 Aug 5; 111(31):E3243-51. PMID: 25049412.
      View in: PubMed
    8. Carone BR, Xu T, Murphy KC, Marinus MG. High incidence of multiple antibiotic resistant cells in cultures of in enterohemorrhagic Escherichia coli O157:H7. Mutat Res. 2014 Jan; 759:1-8. PMID: 24361397.
      View in: PubMed
    9. Ioerger TR, O'Malley T, Liao R, Guinn KM, Hickey MJ, Mohaideen N, Murphy KC, Boshoff HI, Mizrahi V, Rubin EJ, Sassetti CM, Barry CE, Sherman DR, Parish T, Sacchettini JC. Identification of new drug targets and resistance mechanisms in Mycobacterium tuberculosis. PLoS One. 2013; 8(9):e75245. PMID: 24086479.
      View in: PubMed
    10. Murphy KC, Volkert MR. Structural/functional analysis of the human OXR1 protein: identification of exon 8 as the anti-oxidant encoding function. BMC Mol Biol. 2012 Aug 08; 13:26. PMID: 22873401.
      View in: PubMed
    11. Murphy KC. Phage recombinases and their applications. Adv Virus Res. 2012; 83:367-414. PMID: 22748814.
      View in: PubMed
    12. Flockhart AF, Tree JJ, Xu X, Karpiyevich M, McAteer SP, Rosenblum R, Shaw DJ, Low CJ, Best A, Gannon V, Laing C, Murphy KC, Leong JM, Schneiders T, La Ragione R, Gally DL. Identification of a novel prophage regulator in Escherichia coli controlling the expression of type III secretion. Mol Microbiol. 2012 Jan; 83(1):208-23. PMID: 22111928.
      View in: PubMed
    13. Brady MJ, Radhakrishnan P, Liu H, Magoun L, Murphy KC, Mukherjee J, Donohue-Rolfe A, Tzipori S, Leong JM. Enhanced Actin Pedestal Formation by Enterohemorrhagic Escherichia coli O157:H7 Adapted to the Mammalian Host. Front Microbiol. 2011; 2:226. PMID: 22102844.
      View in: PubMed
    14. Tree JJ, Roe AJ, Flockhart A, McAteer SP, Xu X, Shaw D, Mahajan A, Beatson SA, Best A, Lotz S, Woodward MJ, La Ragione R, Murphy KC, Leong JM, Gally DL. Transcriptional regulators of the GAD acid stress island are carried by effector protein-encoding prophages and indirectly control type III secretion in enterohemorrhagic Escherichia coli O157:H7. Mol Microbiol. 2011 Jun; 80(5):1349-65. PMID: 21492263.
      View in: PubMed
    15. Wei JR, Krishnamoorthy V, Murphy K, Kim JH, Schnappinger D, Alber T, Sassetti CM, Rhee KY, Rubin EJ. Depletion of antibiotic targets has widely varying effects on growth. Proc Natl Acad Sci U S A. 2011 Mar 8; 108(10):4176-81. PMID: 21368134.
      View in: PubMed
    16. Murphy KC. Targeted chromosomal gene knockout using PCR fragments. Methods Mol Biol. 2011; 765:27-42. PMID: 21815084.
      View in: PubMed
    17. Murphy KC, Marinus MG. RecA-independent single-stranded DNA oligonucleotide-mediated mutagenesis. F1000 Biol Rep. 2010 Jul 22; 2:56. PMID: 20711416.
      View in: PubMed
    18. Murphy KC, Ritchie JM, Waldor MK, Løbner-Olesen A, Marinus MG. Dam methyltransferase is required for stable lysogeny of the Shiga toxin (Stx2)-encoding bacteriophage 933W of enterohemorrhagic Escherichia coli O157:H7. J Bacteriol. 2008 Jan; 190(1):438-41. PMID: 17981979.
      View in: PubMed
    19. Murphy KC. The lambda Gam protein inhibits RecBCD binding to dsDNA ends. J Mol Biol. 2007 Aug 3; 371(1):19-24. PMID: 17583735.
      View in: PubMed
    20. Campellone KG, Roe AJ, Løbner-Olesen A, Murphy KC, Magoun L, Brady MJ, Donohue-Rolfe A, Tzipori S, Gally DL, Leong JM, Marinus MG. Increased adherence and actin pedestal formation by dam-deficient enterohaemorrhagic Escherichia coli O157:H7. Mol Microbiol. 2007 Mar; 63(5):1468-81. PMID: 17302821.
      View in: PubMed
    21. Savage PJ, Leong JM, Murphy KC. Rapid allelic exchange in enterohemorrhagic Escherichia coli (EHEC) and other E. coli using lambda red recombination. Curr Protoc Microbiol. 2006 Jan; Chapter 5:Unit5A.2. PMID: 18770591.
      View in: PubMed
    22. Murphy KC, Campellone KG. Lambda Red-mediated recombinogenic engineering of enterohemorrhagic and enteropathogenic E. coli. BMC Mol Biol. 2003 Dec 13; 4:11. PMID: 14672541.
      View in: PubMed
    23. Loh T, Murphy KC, Marinus MG. Mutational analysis of the MutH protein from Escherichia coli. J Biol Chem. 2001 Apr 13; 276(15):12113-9. PMID: 11124943.
      View in: PubMed
    24. Murphy KC, Campellone KG, Poteete AR. PCR-mediated gene replacement in Escherichia coli. Gene. 2000 Apr 4; 246(1-2):321-30. PMID: 10767554.
      View in: PubMed
    25. Murphy KC. Use of bacteriophage lambda recombination functions to promote gene replacement in Escherichia coli. J Bacteriol. 1998 Apr; 180(8):2063-71. PMID: 9555887.
      View in: PubMed
    26. Murphy KC. Biochemical characterization of P22 phage-modified Escherichia coli RecBCD enzyme. J Biol Chem. 1994 Sep 9; 269(36):22507-16. PMID: 8077199.
      View in: PubMed
    27. Murphy KC, Lewis LJ. Properties of Escherichia coli expressing bacteriophage P22 Abc (anti-RecBCD) proteins, including inhibition of Chi activity. J Bacteriol. 1993 Mar; 175(6):1756-66. PMID: 8383665.
      View in: PubMed
    28. Murphy KC. Lambda Gam protein inhibits the helicase and chi-stimulated recombination activities of Escherichia coli RecBCD enzyme. J Bacteriol. 1991 Sep; 173(18):5808-21. PMID: 1653221.
      View in: PubMed
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