Design, Synthesis and Evaluation of Triazole‐Pyrimidine Analogues as SecA Inhibitors

Abstract

SecA, a key component of bacterial Sec-dependent secretion pathway, is an attractive target for novel antimicrobial development. Through a combination of virtual screening and experimental exploration of surrounding chemical space, we identified a hit bistriazole SecA inhibitor, SCA-21, and studied a series of analogs by systematic dissections of the core scaffold. Evaluation of these analogs allowed us to establish an initial SAR in SecA inhibition. The best compounds in this group have potent inhibition activity of SecA-dependent protein-conducting channel activity and protein translocation activity at low to sub-μM concentrations. They also have MIC values against various strains of bacteria that are correlated with the SecA and protein translocation inhibition data. These compounds are effective against methicillin-resistant Staplylococcus aureus strains with various levels of efflux pumps, indicating the ability to null the effect of multiple-drug resistance with SecA inhibitors. Results from studies of drug affinity responsive target stability and protein pull-down assays are consistent with SecA as a target for these compounds.

Using Chemical Probes to Assess the Feasibility of Targeting SecA for Developing Antimicrobial Agents against Gram‐Negative Bacteria

“…Earlier, we have noticed that these classes do not bind directly to or inside the ATP containing pockets in chains A and B, but lie close to and outside the ATP pocket, (SCA-50, and SCA-15), [13, 23a] whereas SCA-107 binds on the interface of chains A and B. [9] Such observations are consistent with the kinetic data obtained showing non-competitive inhibition with ATP (Fig. 3).…”

“…Our recent publications detail the discussion of the various assays available and their utilities. [3a, 7b, 9] In particular, we have developed two very sensitive assays to evaluate the functions of SecA in the lipid-environments. [18a, 24] Both are reconstituted SecA-only liposomes systems specific for SecA functions in lipid environment for ion channel activity and protein translocation activity [SFig.…”

“…[13] Its smaller analogues such as SCA-50 are more selective. [7a, 13] Kinetic studies using the channel activity of SecA-only liposomes assayed in the oocytes (Figure 3) showed that all 3 classes of inhibitors Class A SCA-50, Class B SCA-15, and Class C SCA-107 ( [27] used here for comparison) are non-competitive against ATP for EcSecA, PaSecA or StSecA (Figure 3). Thus major inhibitory effects in the membrane may not be on the high-affinity ATPase active site directly (Table 5C); rather, the inhibition is probably allosteric in a membrane lipid environment (SFig.…”

“…[3a, 7a, 13, 23a, 27] All SecA inhibitors were dissolved in DMSO with a final DMSO concentration of 2% in all cultures or in all biochemical assays, which has no effect on cell growth or assays. Polymyxin derivative NAB7061 and PMBN were from Northern Antibiotics Ltd., Helsinki, Finland and Sigma-Aldrich, respectively, and used as received.…”

“…[3a, 5] A recent paper reported a low μM inhibitor against citrus pathogen , Candidatus Liberibacter asiaticus. [6] The three structural classes of small molecule SecA inhibitors (Figure 1) that we developed include Rose Bengal and its Class A analogues, [7] Class B pyrimidine analogues, [8] and Class C triazole-pyrimidine analogues [9] with the best having IC 50 and/or MIC values in the high nM range (Tables 1 and 2). [7b] …”