Assuntos
Clorometilcetonas de Aminoácidos/farmacologia , Retículo Endoplasmático/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Peptídeo-N4-(N-acetil-beta-glucosaminil) Asparagina Amidase/antagonistas & inibidores , Sequência de Aminoácidos , Animais , Sítios de Ligação/genética , Inibidores de Caspase , Retículo Endoplasmático/metabolismo , Humanos , Camundongos , Dados de Sequência Molecular , Mapeamento de Peptídeos , Peptídeo-N4-(N-acetil-beta-glucosaminil) Asparagina Amidase/química , Peptídeo-N4-(N-acetil-beta-glucosaminil) Asparagina Amidase/genética , Peptídeo-N4-(N-acetil-beta-glucosaminil) Asparagina Amidase/metabolismo , Saccharomyces cerevisiae/efeitos dos fármacos , Saccharomyces cerevisiae/enzimologia , Homologia de Sequência de Aminoácidos , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por MatrizRESUMO
The multidrug efflux complex AcrAB-TolC confers intrinsic drug resistance in Escherichia coli by pumping antibiotics out of the cell. We determined a low-resolution (20 A) structure of AcrA, the periplasmic component, by electron crystallography. Expressed with a His-tag at its carboxyl-terminus, the protein bound to lipid layers containing the nickel-chelating phospholipid DOGS-NTA. Under the lipid layers, AcrA crystallized in layer group P2(1)22, with a unit cell size of 157 by 95 A and a thickness of about 100 A. The four asymmetric units in the unit cell are organized into what appears to be two rings, each with a central opening of 30 A in diameter. Within each ring, the density can be interpreted as following a pseudo-helical path, approximately 210 A long. This length matches that of monomeric AcrA in solution, previously estimated by light scattering and hydrodynamic measurements. On one side the density has a tubular shape, with a thickness of about 25 A, while on the other side the densities of the upper and lower parts of the pseudo-helical path are fused into a shield.
