Mechanism of Coupling of Transport to Hydrolysis in Bacterial ATP-Binding Cassette Transporters

Studies of periplasmic binding protein-dependent transporters date to the 1960s when it was realized that the transport of certain solutes was inhibited following an osmotic shock that released the contents of the periplasm (8, 80).The ATP-binding cassette (ABC) superfamily ( 52) was defined in 1986 when homology was detected between a binding protein-dependent transporter and a multidrug efflux pump cloned from human cancer cells (21,42).These proteins, also known as traffic ATPases (4), number 80 in the Escherichia coli genome (13, 23) and 48 in the human genome where many have been linked to human disease (29).In addition to the periplasmic binding protein-dependent transporters that mediate uptake, bacterial cells also contain ABC transporters lacking a binding protein that mediate efflux of compounds such as lipopolysaccharides (15,56,95, 133), capsular polysaccharides (87, 90), antimicrobial agents (7, 98, 126), and toxins (35, 47, 57).A few ABC proteins also lack a transmembrane region and use the same architecture to perform alternative functions such as DNA repair (34).A typical ABC transporter has four domains or subunits, two of which are hydrophobic and are predicted to span the membrane multiple times in an alpha-helical conformation and two of which bind nucleotide and are exposed to the cytoplasm.This domain architecture is clearly established in the recent structure of MsbA (20), an ABC transporter that mediates the export of lipid and lipid A in E. coli (32, 133).MsbA is a homodimer, and the intracellular loops between transmembrane-spanning helices of each monomer unit constitute a novel intracellular alpha-helical domain that physically separates the nucleotide-binding domain from the transmembrane domain.The nucleotide-binding domains and subunits share considerable sequence homology across the entire family (45) and assume a similar three-dimensional fold that consists of a core nucleotide-binding subdomain that is common to other ATPases and an alpha-helical subdomain that is specific to ABC proteins (50).The nucleotide-binding subdomain contains the canonical Walker A and B motifs (128) that are involved in nucleotide binding, while the helical subdomain contains the ABC family signature or LSGGQ motif the function of which is still debatable (31,48,53).This minireview summarizes recent work using vanadate to stabilize the transition state for ATP hydrolysis that has provided new insight into the mechanism of action of this family of proteins.