Crystallization and preliminary X-ray crystallographic analysis of MacA from Actinobacillus actinomycetemcomitans.

Abstract

Periplasmic membrane-fusion proteins (MFPs) are an essential component of the multidrug efflux pump in Gram-negative bacteria. They play a crucial role in bridging the outer membrane porin TolC and two distinct types of inner membrane transporters. The MFP MacA bridges the inner membrane ABC-type multidrug transporter MacB and the outer membrane porin TolC. MacA from the pathogenic bacterium Actinobacillus actinomycetemcomitans was expressed in Escherichia coli B834 (DE3) and the recombinant protein was purified using Ni-NTA affinity, Q anion-exchange and gel-filtration chromatography. The purified MacA protein was crystallized using the vapour-diffusion method. A MAD diffraction data set was collected to a resolution of 3.0 A at 100 K. The crystal belongs to space group P622, with unit-cell parameters a = b = 109.2, c = 255.4 A, alpha = beta = 90, gamma = 120 degrees , and contains one molecule in the asymmetric unit.