Crucial enzymes involved in nucleic acid metabolism and cellular proliferation are targeted for inhibition by synthetic carbohydrate and nucleoside analogues. Living organisms, from bacteria to humans, use S-adenosylmethionine (SAM) as the methyl donor. The byproduct of these reactions is S-adenosylhomocysteine (AdoHcy). Both prokaryotes and eukaryotes have developed detoxification mechanisms to eliminate AdoHcy. Eukaryotes contain an AdoHcy hydrolase, which directly hydrolyzes AdoHcy into homocysteine (Hcy) and adenosine. However, the majority of bacterial species employ a different, two-step mechanism to remove AdoHcy. First, the adenine base of AdoHcy is removed by a 5'- methylthioadenosine/ AdoHcy nucleosidase (MTAN) to produce S-ribosylhomocysteine (SRH). Next, the thioether bond in SRH is cleaved by an S-ribosylhomocysteinase (LuxS enzyme) to produce Hcy and 4,5-dihydroxy-2,3-pentanedione (DPD), important precursors to quorum-sensing-regulated auto-inducers. We are interested in probing substrate similarities and mechanistic differences between AdoHcy hydrolase and LuxS enzymes. The major objective is to prepare modified SRH analogues lacking an enolizable hydroxyl group at carbon-2 or 3. These competitive substrates should act as mechanism-based inhibitors of LuxS and impede the production of autoinducer of type 2, and disrupt quorum sensing and cell-cell communication in bacteria. Ribonucleotide reductases (RNRs) are enzymes that execute 2'-deoxygenation of ribonucleotides and present attractive chemotherapeutic targets for intervention with replication of cancer cells. Our objective is to synthesize a series of 3'-azido-3'-deoxynucleosides bearing a thiol or vicinal dithiol substituent attached to C2' or C5' and perform biomimetic simulation of the reactions postulated to occur during inhibition of RNR by 2'-azido-2'-deoxynucleotides. To expand our findings from the current award, we plan to explore application of organogermanes toward Pd-catalyzed cross-coupling reactions. Relevance to Public Health: Understanding the mechanism of action of the targeted enzymes is essential to pursue the rational design of new mechanism-based inhibitor drugs. Inhibitors of LuxS enzyme should affect quorum sensing regulated processes in bacterial cell-cell communication, which will allow interventions based on virulence, antibiotic production, and biofilm formation.
