How to Find a Fragment: Methods for Screening and Validation in Fragment-Based Drug Discovery.

Fragment-based drug discovery (FBDD) is a crucial strategy for developing new drugs that have been applied to diverse targets, from neglected infectious diseases to cancer. With at least seven drugs already launched to the market, this approach has gained interest in both academics and industry in the last 20 years. FBDD relies on screening small libraries with about 1000-2000 compounds of low molecular weight (about 300 Da) using several biophysical methods. Because of the reduced size of the compounds, the chemical space and diversity can be better explored than large libraries used in high throughput screenings. This review summarises the most common biophysical techniques used in fragment screening and orthogonal validation. We also explore the advantages and drawbacks of the different biophysical techniques and examples of applications and strategies.

Structural Basis for the Interaction and Processing of ß-Lactam Antibiotics by l,d-Transpeptidase 3 (LdtMt3) from Mycobacterium tuberculosis.

Targeting Mycobacterium tuberculosis peptidoglycans with ß-lactam antibiotics represents a strategy to address increasing resistance to antitubercular drugs. ß-Lactams inhibit peptidoglycan synthases such as l,d-transpeptidases, a group of carbapenem-sensitive enzymes that stabilize peptidoglycans through 3 → 3 cross-links. M. tuberculosis encodes five l,d-transpeptidases (LdtMt1-5), of which LdtMt3 is one of the less understood. Herein, we structurally characterized the apo and faropenem-acylated forms of LdtMt3 at 1.3 and 1.8 Å resolution, respectively. These structures revealed a fold and catalytic diad similar to those of other LdtsMt enzymes, supporting its involvement in transpeptidation reactions despite divergences in active site size and charges. The LdtMt3-faropenem structure indicated that faropenem is degraded after Cys-246 acylation, and possibly only a ß-OH-butyrate or an acetyl group (C2H3O) covalently attached to the enzyme remains, an observation that strongly supports the notion that LdtMt3 is inactivated by ß-lactams. Docking simulations with intact ß-lactams predicted key LdtMt3 residues that interact with these antibiotics. We also characterized the heat of acylation involved in the binding and reaction of LdtMt3 for ten ß-lactams belonging to four different classes, and imipenem had the highest inactivation constant. This work provides key insights into the structure, binding mechanisms, and degradation of ß-lactams by LdtMt3, which may be useful for the development of additional ß-lactams with potential antitubercular activity.