Mechanism and Inhibition for LuxS: A Biodefense Target

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DESCRIPTION (provided by applicant): Bacterial infections, particularly those emerging with antibiotic resistance, pose an alarming threat to public health. Our long-term objective is to identify, characterize and validate new antibacterial targets. Traditional antibiotics act by killing or inhibiting bacteria, hence inducing antibiotic resistance. It is therefore imperative to explore alternative or complementary approaches. In the past few years, the ubiquitous bacterial enzyme LuxS has been found to play diverse and pivotal roles in bacterial quorum sensing, virulence regulation, toxin secretion and biofilm formation. In addition, central metabolic roles for LuxS are also proposed. This enzyme is found in Category A pathogens, including B. anthracis and Yersinia pestis; and in Category B pathogens, including Vibrio cholerae, Salmonella and diarrheagenic E. coil. Absent in humans, LuxS is an attractive target for anti-infective agent development. The enzymatic mechanism of LuxS remains elusive. Based on our preliminary studies, we propose that LuxS possesses functions of both an aldose-ketose isomerase and a lyase. The dual function of LuxS is mechanistically intriguing. Our proposed mechanism involves an initial aldose-ketose isomerization to generate a ketone at the C3 position on the carbohydrate moiety, and a final beta-elimination to cleave the C-S bond in S-ribosylhomocysteine. Our Specific Aim 1 is to chemically synthesize the proposed intermediates and their analogs, and test them as LuxS substrates or inhibitors. We will also attempt to trap or directly observe the proposed intermediates. Our Specific Aims 2 and 3 are to investigate the catalytic roles of Glu57 and Cys84 in B. subtilis LuxS by mutagenesis and chemical rescue, and the biological relevance of Cys84 oxidation. Lastly, we plan to design, synthesize and test mechanism-based inhibitors for LuxS, particularly those interacting with the active site zinc ion. We will also investigate how halogenated furanones, a group of natural antibacterial agents, inactivates LuxS, particularly for the V. cholerae enzyme. Additionally, we will test the effects of LuxS inhibitors on quorum sensing, biofilm formation and related bacterial physiology.