Design and synthesis of trans-2-substituted-cyclopropane-1-carboxylic acids as the first non-natural small molecule inhibitors of O-acetylserine sulfhydrylase

Abstract

O-Acetylserine sulfhydrylase (isoform A, OASS-A) is a pyridoxal-5 0 -phosphate-dependent enzyme responsible for cysteine biosynthesis in many pathological microorganisms. It is proposed that inhibition of OASS-A could represent a novel strategy to overcome bacterial resistance to antibiotics. A class of 2-substituted-cyclopropane-1-carboxylic acids was synthesized, based on structural determinants grasped by analyzing a group of synthetic pentapeptides known to efficiently bind OASS-A from Haemophilus influenzae (HiOASS-A). The cyclopropane derivatives were submitted to a binding affinity assay with HiOASS-A and three of them, with K diss in the low micromolar range, showed higher affinity than the most active synthetic pentapeptide. Thus, in this communication we report the first example of potent non-natural small molecule inhibitors of HiOASS-A. In addition, a molecular modelling study suggested a possible inhibition mechanism, through which the new cyclopropane ligands block HiOASS-A. Noteworthily, the novel, small-sized, non-peptidic inhibitors retain the structural motifs of the bulky peptides, which are relevant for the enzyme inhibition.

Cyclopropane-1,2-dicarboxylic acids as new tools for the biophysical investigation ofO-acetylserine sulfhydrylases by fluorimetric methods and saturation transfer difference (STD) NMR

“…On a similar vein, when the ester moiety and the acid are in trans stereo-relationship, the affinity is higher than that measured on the cis stereoisomer. In a more comprehensive picture (Figure 2), this small set of molecules has corroborated the findings partially stemmed also from our previous studies 25,27 : assuming the cyclopropanecarboxylic acid as the pharmacophore, substituents at C-2 showing a hydrophilic character must be kept in a cis stereo-relationship with the C-1 carboxylic group, whereas less hydrophilic substituents must maintain a trans stereo-relationship.…”

“…Since for the most active compounds the main structural motif (the 2-phenylcyclopropylcarboxylic moiety) is maintained, it can be reasonably assumed that the binding mode of these derivatives is similar to that already reported in our previous works [25][26][27] . To ascertain this hypothesis, and to correlate the data obtained from the STD-NMR experiments and the biochemical evaluation, a computational analysis of the binding mode of compound 23 with StOASS-A was performed.…”

“…In order to further refine the structure-activity relationships (SAR) around this class of derivatives, we herein report the design and synthesis of a series of cyclopropane-1,2-dicarboxylic acids variously functionalized. Although the binding properties of the previously reported analogs [24][25][26] could not be improved, the set of compounds herein presented provides interesting insight into the mode of binding of small molecules to OASS enzymes. In particular, saturation transfer difference NMR (STD-NMR) was used to further characterize the molecule/enzyme interactions for both OASS-A and B. Interestingly, most of the compounds induce a several fold increase in fluorescence emission of the pyridoxal 5 0 -phosphate (PLP) coenzyme upon binding to either OASS-A or OASS-B, likely due to changes in the polarity of the active-site microenvironment.…”

“…The main chemical feature was represented by the presence of a carboxylic acid moiety linked through a cyclopropane spacer, to a hydrophobic side chain in a trans configuration. These structures were rationally designed based on the Haemophilus influenzae (H. influenzae) SAT (HiSAT) native Cterminal pentapeptide MNLNI, which is the natural inhibitor of OASS, and the substituent at position C-2 was represented by a small alkenyl chain, as in the case of compound 29 ( Figure 1) 25 . In a second round of optimization, the alkenyl chain was embodied into a phenyl ring 30 , maintaining the trans stereorelationship with the carboxylic moiety.…”

“…The rational design of the first inhibitors was based on the structure of SAT, that physiologically inhibits OASS activity upon formation of the cysteine synthase bioenzyme complex 22,23 ; in particular, it was considered that the carboxylic moiety of Ile267 of SAT is essential for the interaction between the two proteins [22][23][24] . Therefore, combining a structure-based with a ligand-based drug design approach, we planned and synthesized a series of cyclopropanecarboxylic acid derivatives that bind to both OASS isoforms at nanomolar concentrations [25][26][27] . In order to further refine the structure-activity relationships (SAR) around this class of derivatives, we herein report the design and synthesis of a series of cyclopropane-1,2-dicarboxylic acids variously functionalized.…”