Biochemical characterization of phosphoserine phosphatase SerB2 from Mycobacterium marinum.

SerB2 is an essential phosphoserine phosphatase (PSP) that has been shown to be involved in Mycobacterium tuberculosis (Mtb) immune evasion mechanisms, and a drug target for the development of new antitubercular agents. A highly similar (91.0%) orthologous enzyme exists in the surrogate organism Mycobacterium marinum (Mma) and could have acquired similar properties. By homology modeling, we show that the two PSPs are expected to exhibit almost identical architectures. MmaSerB2 folds into a homodimer formed by two intertwined subunits including two ACT regulatory domains followed by a catalytic core typical of HAD (haloacid dehalogenase) phosphatases. Their in vitro catalytic properties are closely related as MmaSerB2 also depends on Mg2+ for the dephosphorylation of its substrate, O-phospho-l-serine (PS), and is most active at neutral pH and temperatures around 40 °C. Moreover, an enzyme kinetics study revealed that the enzyme is inhibited by PS as well, but at lower concentrations than MtbSerB2. Substrate inhibition could occur through the binding of PS in the second active site and/or at the ACT domains interface. Finally, previously described beta-carboline MtbSerB2 inhibitors also decrease the phosphatase activity of MmaSerB2. Altogether, these results provide useful information when M.marinum is used as a model to study immune evasion in tuberculosis.

Identification and Repurposing of Trisubstituted Harmine Derivatives as Novel Inhibitors of Mycobacterium tuberculosis Phosphoserine Phosphatase.

Mycobacterium tuberculosis is still the deadliest bacterial pathogen worldwide and the increasing number of multidrug-resistant tuberculosis cases further complicates this global health issue. M. tuberculosis phosphoserine phosphatase SerB2 is a promising target for drug design. Besides being a key essential metabolic enzyme of the pathogen's serine pathway, it appears to be involved in immune evasion mechanisms. In this work, a malachite green-based phosphatase assay has been used to screen 122 compounds from an internal chemolibrary. Trisubstituted harmine derivatives were found among the best hits that inhibited SerB2 activity. Synthesis of an original compound helped to discuss a brief structure activity relationship evaluation. Kinetics experiments showed that the most potent derivatives inhibit the phosphatase in a parabolic competitive fashion with apparent inhibition constants ( K i ) values in the micromolar range. Their interaction modes with the enzyme were investigated through induced fit docking experiments, leading to results consistent with the experimental data. Cellular assays showed that the selected compounds also inhibited M. tuberculosis growth in vitro. Those promising results may provide a basis for the development of new antimycobacterial agents targeting SerB2.

Targeting the Serine Pathway: A Promising Approach against Tuberculosis?

Tuberculosis is still the leading cause of death by a single infectious agent. Effective chemotherapy has been used and improved since the 1950s, but strains resistant to this therapy and most antibacterial drugs on the market are emerging. Only 10 new drugs are in clinical trials, and two of them have already demonstrated resistance. This paper gives an overview of current treatment options against tuberculosis and points out a promising approach of discovering new effective drugs. The serine production pathway is composed of three enzymes (SerA1, SerC and SerB2), which are considered essential for bacterial growth, and all of them are considered as a therapeutic drug target. Their crystal structure are described and essential regulatory domains pointed out. Sequence alignment with similar enzymes in other host would help to identify key residues to target in order to achieve selective inhibition. Currently, only inhibitors of SerB2 are described in the literature. However, inhibitors of human enzymes are discussed, and could be used as a good starting point for a drug discovery program. The aim of this paper is to give some guidance for the design of new hits for every enzyme in this pathway.