Bacterial resistance to l3-lactam antibiotics continues to become more prevalent and more clinically important. A large part of the resistance can be understood and investigated experimentally in terms of the chemistry of the interactions of R-lactam antibiotics with the active sites of two groups of bacterial enzymes, the Illactamases on one hand, which catalyze the hydrolysis of the antibiotics, and the D-alanyl-D-alanine transpeptidase/carboxypeptidases on the other, which catalyze the synthesis and maintenance of the peptide cross-links of bacterial cell walls, and which are inhibited by 13-lactam antibiotics. There is now good reason to believe that all of these l-lactam binding sites have much in common. An understanding of the structure and function of these sites and of the relationship between them is fundamental to future antibiotics design - both 1-lactam and otherwise. The object of the proposed research is to explore further the chemical functionality and the substrate binding properties of a series of these active sites, using a number of modified substrates, novel inhibitors and potential effedtors. Particular focus will be on the development of novel inhibitors of the (1lactamases and the extension of these inhibitors to DD-peptidases. The specificity of the DD-peptidases for peptidoglycan structural motifs will be examined closely. Crystal structures will be used in conjunction with molecular modeling to interpret the results obtained and apply them to further inhibitor design. In silico and in vitro screening methods will also be used with the OD-peptidases to obtain new lead compounds. These studies will lead to new insight into the chemistry of R-lactamase and transpeptidase active sites, and thus to new directions in antibiotic design. E Gyp 0-. ... G'O --a tail ... 'Fn """"""""-'a'- .N. >.t (Nn Nn. om' m'? """"""""L"""""""" O-- Oil 0.0)-7-.o 9,' -_p >'7 ='L

Public Health Relevance

Relevance Bacteria become more and more resistant to the currently used antibiotics. This entails a huge cost in both health resources and human life. This project involves a study of the enzymes that control the effectiveness of the -lactam antibiotics. The results will provide information aiding new antibiotic development.

National Institute of Health (NIH)
National Institute of Allergy and Infectious Diseases (NIAID)
Research Project (R01)
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Macromolecular Structure and Function E Study Section (MSFE)
Program Officer
Huntley, Clayton C
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Wesleyan University
Schools of Arts and Sciences
United States
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Nemmara, Venkatesh V; Nicholas, Robert A; Pratt, R F (2016) Synthesis and Kinetic Analysis of Two Conformationally Restricted Peptide Substrates of Escherichia coli Penicillin-Binding Protein 5. Biochemistry 55:4065-76
Tilvawala, Ronak; Cammarata, Michael; Adediran, S A et al. (2015) A New Covalent Inhibitor of Class C ?-Lactamases Reveals Extended Active Site Specificity. Biochemistry 54:7375-84
Dzhekieva, Liudmila; Adediran, S A; Pratt, R F (2014) Interactions of ""bora-penicilloates"" with serine ?-lactamases and DD-peptidases. Biochemistry 53:6530-8
Dzhekieva, Liudmila; Adediran, S A; Herman, Raphael et al. (2013) Inhibition of DD-peptidases by a specific trifluoroketone: crystal structure of a complex with the Actinomadura R39 DD-peptidase. Biochemistry 52:2128-38
Tilvawala, Ronak; Pratt, R F (2013) Kinetics of action of a two-stage pro-inhibitor of serine ?-lactamases. Biochemistry 52:7060-70
Tilvawala, Ronak; Pratt, R F (2013) Covalent inhibition of serine ?-lactamases by novel hydroxamic acid derivatives. Biochemistry 52:3712-20
Nemmara, Venkatesh V; Adediran, S A; Dave, Kinjal et al. (2013) Dual substrate specificity of Bacillus subtilis PBP4a. Biochemistry 52:2627-37
Dzhekieva, Liudmila; Kumar, Ish; Pratt, R F (2012) Inhibition of bacterial DD-peptidases (penicillin-binding proteins) in membranes and in vivo by peptidoglycan-mimetic boronic acids. Biochemistry 51:2804-11
Adediran, S A; Lin, G; Pelto, R B et al. (2012) Crossover inhibition as an indicator of convergent evolution of enzyme mechanisms: a ?-lactamase and a N-terminal nucleophile hydrolase. FEBS Lett 586:4186-9
Pelto, Ryan B; Pratt, R F (2012) Kinetics and stereochemistry of hydrolysis of an N-(phenylacetyl)-*-hydroxyglycine ester catalyzed by serine *-lactamases and DD-peptidases. Org Biomol Chem 10:7356-62

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