DOMAIN ORGANIZATION OF MURINE MDR1B P-GLYCOPROTEIN - THE CYTOPLASMIC LINKER REGION IS A POTENTIAL DIMERIZATION DOMAIN

P-glycoprotein is an integral membrane protein that functions as a cyt otoxic drug-efflux pump. Studies suggest that the transporter exists i n the membrane as a dimer and possibly higher order structures. We rep ort the bacterial expression of the linker region (amino acids 621-688 ) of murine mdr1b P-glycoprotein and demonstrate that this region, whi ch serves to link the two homologous halves of the transporter, has th e potential to serve as a dimerization domain. The recombinant peptide (8742 daltons) eluted from a gel filtration column at a position corr esponding to a dimer (i.e. 17,500 daltons). A dimer:monomer equilibriu m, as a function of peptide concentration, was confirmed by large zone gel filtration chromatography. The dissociation constant (K-D) for th e dimer:monomer equilibrium was 350 nM. It was possible to dissociate the dimer by low pH (3.0) or high ionic strength (2.5 M NaCl). Dimeriz ation was not affected by an alkaline pH of 10 or 5 mM EDTA. Studies w ith a truncated linker peptide indicated that the N-terminal 48 amino acids were sufficient for dimerization. (C) 1997 Academic Press