Methicillin-resistant strains of Staphylococcus aureus are a major clinical problem. They are multiple drug resistant, but ineffectiveness of penicillins and beta-lactam antibiotics is the real problem, as these are drug of choice to treat staphylococcal infections. The objective of this research is to further knowledge of mechanisms of methicillin resistance. Resistance is determined by several proteins that interact with penicillin. The interactions among these proteins are critical, but poorly understood. Knowledge of these interrelationships may lead to new drug discovery and new and more effective approaches to therapy./ Resistance is mainly due to production a novel low affinity penicillin bind protein, PBP 2a, a well wall synthetic enzyme. PBP 2a seems to substitute for all other PBPs. mecA, the gene encoding PBP 2a, is regulated by the same regulatory genes that control production of inducible beta-lactamase. Another type of penicillin interactive protein, a penicillin sensory signal transducer BlaR1, signals the cell to express PBP 2a and beta-lactamase, which together mediate all beta-lactam resistance in staphylococci. BlaR1 appears to be a PBP fused to an intracellular Zn++ metalloprotease, and as such may represent a completely new type of transmembrane signaling system. There are three aims.
Aim1. To determine the intracellular pathway by which penicillin binding to BlaR1 signals induction of beta- lactamase and PBP 2a. The effect of specific mutations in BlaR1 on signaling will be determined to prove whether or not Blar1 is a metalloprotease. Putative consensus motifs of this superfamily of proteins will be targeted. The relationship between BlaR1 activation and proteolysis of BlaI, the repressor of the beta-lactamase regulon, will be defined.
Aim 2. To identify PBPs, structural determinants, and other elements that interfere with PBP 2a mediated resistance. Effects of PBP deletion and mutations on PBP 2a mediated resistance will test whether PBP 2a can substitute for other PBPs and where essential functions reside within the molecule. The curious phenomenon of negative selection for expression of PBP 2a that we observed in mec naive cells also will be examined.
Aim 3. To determine when during the cell cycle PBPs are expressed and where they are localized. An electron microscopic method for immunolocalization of specific myc-targeted PBPs in the cell will be developed. To augment information about where PBPs localize, when they are expressed during the cell cycle will be determined by Northern blotting.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI046610-02
Application #
6349926
Study Section
Bacteriology and Mycology Subcommittee 2 (BM)
Project Start
2000-02-01
Project End
2004-01-31
Budget Start
2001-02-01
Budget End
2002-01-31
Support Year
2
Fiscal Year
2001
Total Cost
$312,302
Indirect Cost
Name
University of California San Francisco
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
073133571
City
San Francisco
State
CA
Country
United States
Zip Code
94143
Siboo, Ian R; Chambers, Henry F; Sullam, Paul M (2005) Role of SraP, a Serine-Rich Surface Protein of Staphylococcus aureus, in binding to human platelets. Infect Immun 73:2273-80
Katayama, Yuki; Robinson, D Ashley; Enright, Mark C et al. (2005) Genetic background affects stability of mecA in Staphylococcus aureus. J Clin Microbiol 43:2380-3
Young, David M; Harris, Hobart W; Charlebois, Edwin D et al. (2004) An epidemic of methicillin-resistant Staphylococcus aureus soft tissue infections among medically underserved patients. Arch Surg 139:947-51; discussion 951-3
Katayama, Yuki; Zhang, Hong-Zhong; Chambers, Henry F (2004) PBP 2a mutations producing very-high-level resistance to beta-lactams. Antimicrob Agents Chemother 48:453-9
Chambers, Henry F (2003) Solving staphylococcal resistance to beta-lactams. Trends Microbiol 11:145-8
Katayama, Yuki; Zhang, Hong-Zhong; Hong, Dong et al. (2003) Jumping the barrier to beta-lactam resistance in Staphylococcus aureus. J Bacteriol 185:5465-72
Katayama, Yuki; Zhang, Hong-Zhong; Chambers, Henry F (2003) Effect of disruption of Staphylococcus aureus PBP4 gene on resistance to beta-lactam antibiotics. Microb Drug Resist 9:329-36
Winston, Lisa G; Bangsberg, David R; Chambers 3rd, Henry F et al. (2002) Epidemiology of vancomycin-resistant Enterococcus faecium under a selective isolation policy at an urban county hospital. Am J Infect Control 30:400-6
Charlebois, Edwin D; Bangsberg, David R; Moss, Nicholas J et al. (2002) Population-based community prevalence of methicillin-resistant Staphylococcus aureus in the urban poor of San Francisco. Clin Infect Dis 34:425-33
Chambers, H F (2001) The changing epidemiology of Staphylococcus aureus? Emerg Infect Dis 7:178-82

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