Abstract

The function of many genes cannot be deduced from sequence similiarity, and biochemical methods are usually required. Whole genome sequences can be thought of as not only a set of genes but also collections of functional domains. These domains can be studied by affinity methods whereby identification of the ligand can provide information on biochemical function. To take advantage of this method, one must express all functional domains in a form suitable for affinity studies. Phage display technology provides a means for accomplishing this. Phage display libraries that include all of the open reading frames (ORFs) of Treponema pallidum will be constructed. T. pallidum is the causative agent of syphilis. The complete genome sequence of this organism has recently been completed. Several features of T. pallidum make it an excellent system on which to develop and test genomic phage display technology. First, with a size of 1 million base pairs, the genome is one of the smallest known. Second, there are a total of 1041 open reading frames which makes it feasible to systematically construct libraries containing each ORF in a relatively short period of time. Finally, little is known of the biology or pathogensis of this organism because a continuous culture system is not available. This severely limits the experimental options for study of the organism. Therefore, new approaches are needed to understand gene function in T. pallidum. Two types of phage display libraries will be constructed. One type will be constructed by systematically inserting each of the 1041 ORFs of T. pallidum into a phage display vector. The second library will be constructed by shotgun cloning of randomly fragmented T. pallidum genomic DNA. The libraries will be used to identify ORFs that are involved in attachment of T. pallidum to fibronectin and host cells. In addition, the libraries will be used to identify T. pallidum DNA binding proteins and locate their binding sites. This information can be used to discover T. pallidum transcriptonal regulatory networks. The work will also serve as a model system for the use of genomic phage display as a tool for functional genomics.

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI045842-02
Application #
6170964
Study Section
Special Emphasis Panel (ZRG1-BM-1 (04))
Program Officer
Quackenbush, Robert L
Project Start
1999-07-15
Project End
2002-06-30
Budget Start
2000-07-01
Budget End
2001-06-30
Support Year
2
Fiscal Year
2000
Total Cost
$216,606
Indirect Cost

  Institution

Name
Baylor College of Medicine
Department
Microbiology/Immun/Virology
Type
Schools of Medicine
DUNS #
074615394
City
Houston
State
TX
Country
United States
Zip Code
77030

  Publications

Titz, Bjorn; Rajagopala, Seesandra V; Goll, Johannes et al. (2008) The binary protein interactome of Treponema pallidum--the syphilis spirochete. PLoS One 3:e2292
Brinkman, Mary Beth; McGill, Melanie A; Pettersson, Jonas et al. (2008) A novel Treponema pallidum antigen, TP0136, is an outer membrane protein that binds human fibronectin. Infect Immun 76:1848-57
Matejkova, Petra; Strouhal, Michal; Smajs, David et al. (2008) Complete genome sequence of Treponema pallidum ssp. pallidum strain SS14 determined with oligonucleotide arrays. BMC Microbiol 8:76
Brinkman, Mary Beth; McKevitt, Matthew; McLoughlin, Melanie et al. (2006) Reactivity of antibodies from syphilis patients to a protein array representing the Treponema pallidum proteome. J Clin Microbiol 44:888-91
McKevitt, Matthew; Brinkman, Mary Beth; McLoughlin, Melanie et al. (2005) Genome scale identification of Treponema pallidum antigens. Infect Immun 73:4445-50
Smajs, David; McKevitt, Matthew; Howell, Jerrilyn K et al. (2005) Transcriptome of Treponema pallidum: gene expression profile during experimental rabbit infection. J Bacteriol 187:1866-74
McKevitt, Matthew; Patel, Krupa; Smajs, David et al. (2003) Systematic cloning of Treponema pallidum open reading frames for protein expression and antigen discovery. Genome Res 13:1665-74
Materon, Isabel C; Queenan, Anne Marie; Koehler, Theresa M et al. (2003) Biochemical characterization of beta-lactamases Bla1 and Bla2 from Bacillus anthracis. Antimicrob Agents Chemother 47:2040-2
Majiduddin, Fahd K; Materon, Isabel C; Palzkill, Timothy G (2002) Molecular analysis of beta-lactamase structure and function. Int J Med Microbiol 292:127-37
Hoe, N P; Kordari, P; Cole, R et al. (2000) Human immune response to streptococcal inhibitor of complement, a serotype M1 group A Streptococcus extracellular protein involved in epidemics. J Infect Dis 182:1425-36

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