SERCEB: Polyamine Inhibition a Target for Biodefense in Y. Pestis

Yersinia pestis, the causative agent of bubonic and pneumonic plague, has caused devastating pandemics throughout history and is considered a Class A bioterrorism agent. We have developed compelling evidence that polyamines are critical for Y. pestis. We show that polyamine deficient strains (1) have growth deficiencies that cannot be overcome by transport; (2) have severe deficiencies in generation of a biofilm, which is an important mechanism of bacterial resistance to antibiotics and immune response; and (3) have severe depletion in levels of polyamine. The evidence suggests inhibition of bacterial polyamine biosynthesis as a potential novel strategy for the design of antimicrobial agents. Our overall goal is to understand the function of polyamines in growth and virulence of Y. pestis. The focus of this proposal is to design chemical probes that will inhibit polyamine biosynthesis as potential biodefense agents. Aim 1: To s nthesize a libra 0 ar inine analo es that will be used to screen or ol amine bio nthetic inhibition of ADC and the abilitv to disruPt the formation of biofilm. We will generate a library of arginine analogues. This library will be used in an enzyme-based and biofilm-disruption screen to identity polyamine biosynthetic inhibitors that can disrupt biofilm formation. We will target arginine decarboxylase (ADC). We will also screen for alternative substrates which will be converted into suicide inhibitors using a classical mechanism based inhibition strategy used on pyridoxal-S'-phosphate (PLP) dependent enzymes. Aim 2: To determine the structures of inhibitor complexes of ADC for drug optimization. The structural information of inhibitor complexes will be used in drug optimization. We have cloned expressed, purified, crystallized, collected diffraction data and identified a mercury derivative that will allow us to determine its molecular structure.