Inhibitors Against Metallo-Beta-Lactamases

A. J. VILA

Conferencia; IDWeek, ADVANCING SCIENCE, IMPROVING CARE; 2019

Protein evolution can be describedas a walk on sequence space where a fitness value is assigned to eachparticular sequence. A major challenge to understand thebiological mechanisms of protein evolution is the attempt to correlate genotypewith phenotype and fitness at a molecular level. This is a complex challenge,involving the assessment of biochemical, biophysical and structural features tomany protein variants. Antibiotic resistance mediated by β-lactamasesis an ideal system to study protein evolution, since the chances of a whole organismto survive depend on the availability of a folded, stable and active protein inthe proper cellular compartment (the bacterial periplasm).  Metallo-β-lactamases(MBLs) are Zn(II)-dependent β-lactamases that constitute the latest resistancemechanism of pathogenic and opportunistic bacteria against carbapenems,considered as last resort drugs. Zn(II) binding is critical in the bacterialperiplasm, not only to activate these enzymes and provide resistance, but alsoto stabilize the protein scaffold. This phenomenon is not paralleled by invitro studies. We developed a strategy aimed to correlate the biochemical andbiophysical features in purified enzymes with those in the bacterial periplasm,ultimately leading to the selected phenotype, i.e., resistance to antibiotics. This strategy allows us to dissectthe molecular features that are tailored by accumulating mutations duringevolution. We have applied this approach to invitro evolved protein in the laboratory, as well as to natural allelicvariants selected in clinical strains. This has allowed us to account for theepistatic interactions between mutations at a structural level. Finally, we haveshown that the natural evolutionary landscape of allelic variants of aclinically relevant lactamase (NDM) has been shaped by Zn(II) deprivationconditions as those induced by the host immune response. As a consequence,natural NDM variants with enhanced Zn(II) binding affinity have been selected,overriding the most common evolutionary pressure acting on catalyticefficiency.