IBR   13079
INSTITUTO DE BIOLOGIA MOLECULAR Y CELULAR DE ROSARIO
Unidad Ejecutora - UE
congresos y reuniones científicas
Título:
Architecture of the Staphylococcus aureus beta-lactam Sensor MecR1
Autor/es:
BELLUZO, B.S.; ABRIATA, L.A.; DAL PERARO, M.; LLARRULL, L.I.
Lugar:
San Diego
Reunión:
Congreso; ICAAC/ICC 2015; 2015
Institución organizadora:
American Society for Microbiology
Resumen:
Background: In Methicillin-resistant Staphylococcus aureus (MRSA), the sensor protein MecR1 regulates the expression of PBP2a, a transpeptidase not inhibited by clinical concentrations of β-lactams. MecR1 consists of a periplasmic penicillin-binding domain of known structure, and a transmembrane metallopeptidase domain that remains unamenable to structural studies precluding any molecular insights into the mechanism that triggers resistance upon beta-lactam binding. Methods: We constructed mecR1 C-terminal deletion fusions to egfp that positioned the fluorescent protein between each of the six predicted transmembrane domains, testing each fusion for fluorescence and level of expression. We combined homology modeling in Robetta, I-TASSER and Swiss-Model with coevolution-based constraints from EV-Fold and NMR data on the interactions between both domains to obtain a model of MecR1?s architecture. Molecular dynamics simulations were used to drive assembly and to assess the stability of the resulting complex when embedded in a membrane.Results: A gluzincin fold was consistently assigned to the core of MecR1?s peptidase domain by homology modeling, further supported by analysis of residue coevolution. The modelled transmembrane helix topology of MecR1 was inconsistent with sequence-based predictions, but agreed with our experimental results on the MecR1-EGFP C-terminal fusions. An open-sided chamber was defined that did not reach the outer polar region of the membrane, but was closed by the sensor domain in the full protein. The base of the hydrophilic chamber hosted the zinc site. Conclusions: We put forward a model for full-length MecR1 where the sensor domain loops whose dynamics are affected by beta-lactam binding are located inside a hydrophilic transmembrane chamber defined by the metallopeptidase domain, poised to propagate the perturbation to the zinc-containing gluzincin core. This model gives for the first time a possible molecular route for signal transduction in this class of sensor proteins.