We will continue to study aspects of the biosynthesis, assembly and degradation of the bacteria cell wall, especially in relationship to cell growth and cell division. Emphasis will be placed on studies of Streptococcus faecium ATCC 9790 as a """"""""model system,"""""""" particularly because of its relatively simple shape and mode of division. Studies during the next 5-year grant period will be directed towards studies of the possible roles of two separate and distinct muramidases of SF in surface assembly and division. Thus we propose to: (i) Study several of the properties of the two very unusual autolytic murasidases of SF. Included will be studies of the molecular mechanism of hydrolysis of PGs by muramidase-2 (E- 2). The possibility that E-2 is a transglycosidase is of particular interest, as is the possibility that E-2 is a """"""""two- headed"""""""" type of transglycosidase-transpeptidase, similar to at least two of the penicillin-binding proteins (PBPs) of Escherichia coli. Additionally we will study and compare the binding of and catalytic properties of both E-l and E-2 on a number of different soluble and insoluble substrates. The ultimate goal of these studies is to obtain an insight into the action of and regulation of these two activities in growing and dividing bacteria. (ii) Study the possible in vivo functions of these two, redundant, PG hydrolases in cell wall assembly, surface enlargement and/or cell division. Towards this goal we will obtain and characterize isogenic pairs, one of which will contain a mutation in the structural gene for E-1 or E-2. The genes for E-l and E-2 will be cloned in E. coli and the two genes will be sequenced. (iii) Additional studies will include investigations of the relationship of E-2 to PBPs, and studies of cell wall organization using monoclonal antibodies to epitopes present in the cell wall PG.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI005044-28
Application #
3124219
Study Section
Microbial Physiology and Genetics Subcommittee 2 (MBC)
Project Start
1978-01-01
Project End
1993-03-31
Budget Start
1991-04-01
Budget End
1992-03-31
Support Year
28
Fiscal Year
1991
Total Cost
Indirect Cost
Name
Temple University
Department
Type
Schools of Medicine
DUNS #
City
Philadelphia
State
PA
Country
United States
Zip Code
19122
Mitchell, Caroline; Jennings, Cheryl; Brambilla, Donald et al. (2008) Diminished human immunodeficiency virus type 1 DNA yield from dried blood spots after storage in a humid incubator at 37 degrees C compared to -20 degrees C. J Clin Microbiol 46:2945-9
Shockman, G D (1992) The autolytic ('suicidase') system of Enterococcus hirae: from lysine depletion autolysis to biochemical and molecular studies of the two muramidases of Enterococcus hirae ATCC 9790. FEMS Microbiol Lett 79:261-7
Joris, B; Englebert, S; Chu, C P et al. (1992) Modular design of the Enterococcus hirae muramidase-2 and Streptococcus faecalis autolysin. FEMS Microbiol Lett 70:257-64
Kariyama, R; Shockman, G D (1992) Extracellular and cellular distribution of muramidase-2 and muramidase-1 of Enterococcus hirae ATCC 9790. J Bacteriol 174:3236-41
Chu, C P; Kariyama, R; Daneo-Moore, L et al. (1992) Cloning and sequence analysis of the muramidase-2 gene from Enterococcus hirae. J Bacteriol 174:1619-25
Kariyama, R; Massidda, O; Daneo-Moore, L et al. (1990) Properties of cell wall-associated DD-carboxypeptidase of Enterococcus hirae (Streptococcus faecium) ATCC 9790 extracted with alkali. J Bacteriol 172:3718-24
Dolinger, D L; Daneo-Moore, L; Shockman, G D (1989) The second peptidoglycan hydrolase of Streptococcus faecium ATCC 9790 covalently binds penicillin. J Bacteriol 171:4355-61
Dolinger, D L; Schramm, V L; Shockman, G D (1988) Covalent modification of the beta-1,4-N-acetylmuramoylhydrolase of Streptococcus faecium with 5-mercaptouridine monophosphate. Proc Natl Acad Sci U S A 85:6667-71
Pucci, M J; Hinks, E T; Dicker, D T et al. (1986) Inhibition of beta-lactam antibiotics at two different times in the cell cycle of Streptococcus faecium ATCC 9790. J Bacteriol 165:682-8
Shockman, G D; Kawamura, T; Barrett, J F et al. (1985) The autolytic peptidoglycan hydrolases of Streptococcus faecium. Ann Inst Pasteur Microbiol 136A:63-6