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CBE = Cristalografía y Biología Estructural
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Acta Cryst. (1948) 1, 3-4
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Streptococcus pneumoniae is a leading killer of children and immunocompromised individuals. S. pneumoniae has become increasingly resistant to major antibiotics, and therefore the development of new antibiotic strategies is desperately needed. Targeting bacterial cell division is one such strategy, specifically targeting essential proteins for the synthesis and breakdown of peptidoglycan. Across multiple species of bacteria, the protein FtsX is a cell division protein involved in the regulation of peptidoglycan hydrolases. FtsX represents a large group of ABC-transporter like proteins that function as ‘mechanotransmitters’, proteins that relay signals from inside the cell to the outside. In a collaborative international (Spain, Norway and USA) effort leaded by IQFR and University Indiana Bloomington we present the first structural characterization of the large extracellular loop of FtsX from the human opportunistic pathogen Streptococcus pneumoniae. We show the direct interaction between the peptidoglycan hydrolase PcsB and FtsX, and that this interaction is essential for cell viability. As such, FtsX represents an attractive, conserved target for the development of new classes of antibiotics.
mBio (2019) (doi:10.1128/mBio.02622-18)
Journal of Medicinal Chemistry (2018) (doi:10.1021/acs.jmedchem.8b00088)
The quinazolinones are a new class of antibacterials with in vivo efficacy against methicillin resistant Staphylococcus aureus (MRSA). The quinazolinones target cell-wall biosynthesis and have a unique mechanism of action by binding to the allosteric site of penicillin-binding protein (PBP)2a. The combination of the quinazolinone with the commercial piperacillin-tazobactam showed bactericidal synergy. We demonstrated the efficacy of the triple-drug combination in a mouse MRSA neutropenic thigh-infection model. The proposed mechanism for the synergistic activity in MRSA involves inhibition of the β-lactamase by tazobactam, which protects piperacillin from hydrolysis, which can then inhibit its target PBP2a. Furthermore, the quinazolinone binds to the allosteric site of PBP2a triggering the allosteric response. This leads to the opening of the active site, which in turn binds another molecule of piperacillin. The collective effect is the impairment of cell-wall biosynthesis, with bactericidal consequence. Two crystal structures for complexes of the antibiotics with PBP2a provide support for the proposed mechanism of action.
Antimicrob. Agents Chemother. (2019) In press (doi: 10.1128/AAC.02637-18)
β-lactam antibiotics are currently the most used antibiotics. These antibiotics prevent bacterial cell wall formation, critical for bacterial survive. The cell wall contains the PG which is formed by alternated units of N-acetilglucosamine (NAG) and N-acetilmuramic acid (NAM) with peptide stems attached to NAM. These peptide stems are cross-linked creating a net. β-lactam antibiotics inhibit the transpeptidation process between peptides promoting the accumulation of aberrant long fragments. Pseudomonas aeruginosa attempts to repair this damage by the lytic transglycosilase Slt. In our work we have managed to solve the structure of the enzyme Slt to carry out the study of its catalytic mechanism getting several complexes with analogous of its natural substrate. Slt is able to degrade the PG through both endolytic (in the middle of chain) and exolytic (in one end) cut. Slt can accommodate the PG thanks to a long catalytic throat with up to 10 positions for NAG/NAM units together with certain key residues that interact with the peptide stems. These results disclose the details of bacterial response to the β-lactam antibiotic challenge.
Proceedings of the National Academy of Sciences, PNAS (2018) 115, 4393-4398 (doi:10.1073/pnas.1801298115)
Transpeptidases, members of the penicillin-binding protein (PBP) families, catalyze crosslinking of the bacterial cell wall. This transformation is critical for the survival of bacteria and it is the target of inhibition by β-lactam antibiotics. We report herein our structural insights into catalysis by the essential PBP2x of Streptococcus pneumoniae by disclosing a total of four X-ray structures, two computational models based on the crystal structures and molecular-dynamics simulations. The X-ray structures are for the apo PBP2x, the enzyme modified covalently in the active site by oxacillin (a penicillin antibiotic), the enzyme modified by oxacillin in the presence of a synthetic tetrasaccharide surrogate for the cell-wall peptidoglycan and a non-covalent complex of cefepime (a cephalosporin antibiotic) bound to the active site. A pre-requisite for catalysis by transpeptidases, including PBP2x, is the molecular recognition of nascent peptidoglycan strands, which harbor pentapeptide stems. We disclose that the recognition of nascent peptidoglycan by PBP2x takes place by complexation of one pentapeptide stem at an allosteric site located in the PASTA domains of this enzyme. This binding predisposes the third pentapeptide stem in the same nascent peptidoglycan strand to penetration into the active site for the turnover events. The complexation of the two pentapeptide stems in the same peptidoglycan strand is a recognition motif for the nascent peptidoglycan, critical for the cell-wall crosslinking reaction.
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