Nucleotide-Binding Sites of the Heterodimeric LmrCD ABC-Multidrug Transporter of Lactococcus lactis Are AsymmetricLubelski, J., Merkerk, R. V., Konings, W. N. & Driessen, A. J. M., 17-Jan-2006, In : Biochemistry. 45, 2, p. 648-656 9 p.
Research output: Contribution to journal › Article › Academic › peer-review
LmrCD is a lactococcal, heterodimeric multidrug transporter, which belongs to the ABC superfamily. It consists of two half-transporters, LmrC and LmrD, that are necessary and sufficient for drug extrusion and ATP hydrolysis. LmrCD is asymmetric in terms of the conservation of the functional motifs of the nucleotide-binding domains (NBDs). Important residues of the nucleotide-binding site of LmrC and the C loop of LmrD are not conserved. To investigate the functional importance of the LmrC and LmrD subunits, the putative catalytic base residue adjacent to the Walker B motif of both NBDs were substituted for the respective carboxamides. Our data demonstrate that Glu587 of LmrD is essential for both drug transport and ATPase activity of the LmrCD heterodimer, whereas mutation of Asp495 of LmrC has a less severe effect on the activity of the complex. Structural and/or functional asymmetry is further demonstrated by differential labeling of both subunits by 8-azido-[alpha-P-32]ATP, which, at 4 degrees C, occurs predominantly at LmrC, while aluminiumfluoride (AlFx)-induced trapping of the hydrolyzed nucleotide at 30 degrees C results in an almost exclusive labeling of LmrD. It is concluded that the LmrCD heterodimer contains two structurally and functionally distinct NBDs.
|Number of pages||9|
|Publication status||Published - 17-Jan-2006|
- HUMAN P-GLYCOPROTEIN, RESISTANCE PROTEIN MRP1, ATP HYDROLYSIS, CRYSTAL-STRUCTURE, CATALYTIC CYCLE, ESCHERICHIA-COLI, CASSETTE TRANSPORTERS, CARBOXYLATE RESIDUES, MUTATIONAL ANALYSIS, GLUTAMATE RESIDUES