ABCMdb

ABCG8 p.Arg111Met

Predicted by SNAP2:	A: D (85%), C: D (85%), D: D (95%), E: D (91%), F: D (91%), G: D (91%), H: D (85%), I: D (91%), K: N (78%), L: D (91%), M: D (85%), N: D (91%), P: D (95%), Q: D (85%), S: D (85%), T: D (91%), V: D (91%), W: D (95%), Y: D (91%),
Predicted by PROVEAN:	A: D, C: D, D: D, E: N, F: D, G: D, H: D, I: D, K: N, L: D, M: D, N: D, P: D, Q: N, S: D, T: D, V: D, W: D, Y: D,

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Publications
[hide] Functional asymmetry of nucleotide-binding domains... J Biol Chem. 2006 Feb 17;281(7):4507-16. Epub 2005 Dec 12. Zhang DW, Graf GA, Gerard RD, Cohen JC, Hobbs HH
Functional asymmetry of nucleotide-binding domains in ABCG5 and ABCG8.
J Biol Chem. 2006 Feb 17;281(7):4507-16. Epub 2005 Dec 12., 2006-02-17 [PMID:16352607]

Abstract [show]
The ATP-binding cassette half-transporters ABCG5 (G5) and ABCG8 (G8) promote secretion of neutral sterols into bile, a major pathway for elimination of sterols. Mutations in either ABCG5 or ABCG8 cause sitosterolemia, a recessive disorder characterized by impaired biliary and intestinal sterol secretion, sterol accumulation, and premature atherosclerosis. The mechanism by which the G5G8 heterodimer couples ATP hydrolysis to sterol transport is not known. Here we examined the roles of the Walker A, Walker B, and signature motifs in the nucleotide-binding domains (NBD) of G5 and G8 using recombinant adenoviruses to reconstitute biliary sterol transport in G5G8-deficient mice. Mutant forms of each half-transporter were co-expressed with their wild-type partners. Mutations at crucial residues in the Walker A and Walker B domains of G5 prevented biliary sterol secretion, whereas mutations of the corresponding residues in G8 did not. The opposite result was obtained when mutations were introduced into the signature motif; mutations in the signature domain of G8 prevented sterol transport, but substitution of the corresponding residues in G5 did not. Taken together, these findings indicate that the NBDs of G5 and G8 are not functionally equivalent. The integrity of the canonical NBD formed by the Walker A and Walker B motifs of G5 and the signature motif of G8 is essential for G5G8-mediated sterol transport. In contrast, mutations in key residues of the NBD formed by the Walker A and B motifs of G8 and the signature sequence of G5 did not affect sterol secretion.

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Sentences [show]
No. Sentence Comment
53 For G8, R111M-forward (5Ј-TCA GGC TGC GGG ATG GCC TCA CTA CTC-3Ј), R111M- reverse (5ЈGAG TAG TGA GGC CAT CCC GCA GCC TGA-3Ј), R111K-forward (5Ј-TCA GGC TGC GGG AAG GCC TCA CTA CTC-3Ј), R111K-reverse (5ЈGAG TAG TGA GGC CTT CCC GCA GCC TGA-3Ј), G216D-forward (5Ј-GGG GTG TCC GGG GAT GAG CGC CGA CGA-3Ј), G216D-reverse (5Ј-TCG TCG GCG CTC ATC CCC GGA CAC CCC-3Ј), V214IG215S-forward (5Ј-TAT GTA CGT GGG ATC TCC TCG GGT GAG CGC CGA-3Ј), V214IG215S-reverse (5Ј- TCG GCG CTC ACC CGA GGA GAT CCC ACG TAC ATA-3Ј), E238D-forward (5Ј-CTC ATT CTG GAT GAC CCC ACT TCT GGC-3Ј), E238D-reverse (5Ј-GCC AGA AGT GGG GTC ATC CAG AAT GAG-3Ј), E238Q-forward (5Ј-CTC ATT CTG GAT CAG CCC ACT TCT GGC-3Ј), and E238D-reverse (5Ј-GCC AGA AGT GGG CTG ATC CAG AAT GAG-3Ј). Login to comment
156 In contrast to these results, replacement of the corresponding basic residue in the Walker A motif of G8 with methionine (R111M) or with lysine (R111K) had no effect on either the amount of immunodetectable G5 or G8 or on the ability of G5 or G8 to bind ATP or trap ADP (Fig. 5A). Login to comment
174 Substitution of Arg111 in G8 to methionine or lysine had no adverse effects on biliary cholesterol or plant sterol levels, even when steady state levels of mature G5 and G8 were significantly reduced, as occurred in the mice expressing G8-R111M with wild-type G5 (Fig. 5B). Login to comment
189 Mutations in the Walker A motifs of G5 (K93M and K93R) and G8 (R111M and R111K); effects on protein mass, nucleotide binding and trapping in cultured cells (A) and biliary cholesterol and plant sterol secretion in vivo (B). Login to comment