Inhibition of Mycobacterium tuberculosis Transaminase BioA by Aryl Hydrazines and Hydrazides

Abstract

7,8-diaminopelargonic acid synthase (BioA) of Mycobacterium tuberculosis is a recently validated target for therapeutic intervention in the treatment of tuberculosis (TB). Using biophysical fragment screening and structural characterization of compounds, we have identified a potent aryl hydrazine inhibitor of BioA that reversibly modifies the pyridoxal-5’-phosphate (PLP) cofactor, forming a stable quinonoid. Analogous hydrazides also form covalent adducts that can be observed crystallographically but are incapable of inactivating the enzyme. In the X-ray crystal structures, small molecules induce unexpected conformational remodeling in the substrate binding site. We compare these conformational changes to those induced upon binding of the substrate (7-keto-8-aminopelargonic acid), and characterize the inhibition kinetics and the X-ray crystal structures of BioA with the hydrazine compound and analogs to unveil the mechanism of this reversible covalent modification.

Target-Based Identification of Whole-Cell Active Inhibitors of Biotin Biosynthesis in Mycobacterium tuberculosis

“…However, the chemical instability and highly polar nature of this compound precludes its use in vivo (Shi, et al, 2011). The chemical precedence provided by amiclenomycin in conjunction with our successful fragment-screening campaign supports the vulnerability of Mtb to chemical inhibition of BioA (Dai, et al, 2014; Edfeldt, et al, 2011). …”

“…Three complex structures were obtained including 7-diethylaminocoumarin 15 , pyrrolothiazolidine- S -oxide 18 , and an N -aryl piperazine 14-Cl , which is otherwise identical to hit N -aryl piperazine 14 except the dioxolane ring is replaced with a meta -chloro substituent. Unambiguous electron density affirms that each inhibitor binds in the hydrophobic site adjacent to the PLP co-factor where substrate KAPA also binds as shown in Figure 5 (Dai, et al, 2014). This site exists at the interface of monomers in the BioA heterodimer; inhibitors are in contact with structural components of both monomers, distinguished by color in Figure 5 (panels A-C).…”

“…None of the hits showed activity in the BioD counter-screen (IC 50 > 20 μM). Since BioA is a PLP-dependent aminotransferase and because previous BioA inhibitors have been shown to covalently bind the PLP cofactor, we evaluated all compounds against aspartate transaminase (AST), a ubiquitous and functionally related PLP-dependent enzyme, in order to assess potential enzyme selectivity (AID#743184; Dai, et al, 2014; Sandmark, et al, 2002; Shi, et al, 2011; Zlitni, et al, 2013). All of the hits were inactive against AST suggesting they possess a useful level of selectivity.…”

“…Prior structural studies with Mtb BioA have clearly shown that the active site is highly adaptable in the presence of different bound ligands (Dai, et al, 2014; Dey, et al, 2010; Shi, et al, 2011). This plasticity is not unexpected, given the need for the enzyme to recognize, bind, and catalyze chemical transformations of such diverse substrates as KAPA and SAM.…”

Targeting protein biotinylation enhances tuberculosis chemotherapy

“…We previously reported the design and characterization of potent inhibitors of biotin protein ligase (BPL), the enzyme responsible for covalently ligating biotin onto the ACCs (12)(13)(14)(15)(16)(17)(18)(19). This led to 5′-[N-(d-biotinoyl)sulfamoyl]amino-5′-deoxyadenosine (Bio-AMS), a BPL inhibitor with potent on-target whole-cell activity against drugsensitive and drug-resistant Mtb (17).…”