ABCMdb

ABCB1 p.Lys380Arg

Predicted by SNAP2:	A: N (57%), C: D (53%), D: N (78%), E: N (87%), F: D (63%), G: N (57%), H: N (53%), I: N (66%), L: N (61%), M: N (66%), N: N (82%), P: N (78%), Q: N (72%), R: N (82%), S: N (72%), T: N (87%), V: N (61%), W: D (63%), Y: D (63%),
Predicted by PROVEAN:	A: N, C: D, D: N, E: N, F: D, G: D, H: N, I: D, L: D, M: N, N: N, P: N, Q: N, R: N, S: N, T: N, V: N, W: D, Y: D,

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Publications
[hide] Characterization of YvcC (BmrA), a multidrug ABC t... Biochemistry. 2004 Jun 15;43(23):7491-502. Steinfels E, Orelle C, Fantino JR, Dalmas O, Rigaud JL, Denizot F, Di Pietro A, Jault JM
Characterization of YvcC (BmrA), a multidrug ABC transporter constitutively expressed in Bacillus subtilis.
Biochemistry. 2004 Jun 15;43(23):7491-502., 2004-06-15 [PMID:15182191]

Abstract [show]
The involvement of transporters in multidrug resistance of bacteria is an increasingly challenging problem, and most of the pumps identified so far use the protonmotive gradient as the energy source. A new member of the ATP-binding cassette (ABC) family, known in Bacillus subtilis as YvcC and homologous to each half of mammalian P-glycoprotein and to LmrA of Lactococcus lactis, has been studied here. The yvcC gene was constitutively expressed in B. subtilis throughout its growth, and a knockout mutant showed a lower rate of ethidium efflux than the wild-type strain. Overexpression of yvcC in Escherichia coli allowed the preparation of highly enriched inverted-membrane vesicles that exhibited high transport activities of three fluorescent drugs, namely, Hoechst 33342, doxorubicin, and 7-aminoactinomycin D. After solubilization with n-dodecyl beta-D-maltoside, the hexahistidine-tagged YvcC was purified by a one-step affinity chromatography, and its ability to bind many P-glycoprotein effectors was evidenced by fluorescence spectroscopy experiments. Collectively, these results showed that YvcC is a multidrug ABC transporter functionally active in wild-type B. subtilis, and YvcC was therefore renamed BmrA for Bacillus multidrug resistance ATP. Besides, reconstitution of YvcC into liposomes led to the highest, vanadate-sensitive, ATPase activity reported so far for an ABC transporter. Interestingly, such a high ATP hydrolysis proceeds with a positive cooperativity mechanism, a property only found so far with ABC importers.

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No. Sentence Comment
121 It is shown here that ATP hydrolysis is required to fulfill the Hoechst transport since a YvcC mutant, which was altered in the Walker A-motif (K380R) and therefore deficient in ATP hydrolysis (ATPase activity of the K380R mutant <3% of that of the wild-type protein), was unable to carry out such a transport (Figure 4A). Login to comment
132 When AMPPNP was used instead of ATP, no transport activity was observed with wild-type YvcC, and likewise the YvcC K380R mutant was unable to carry out drug transport in the presence of ATP. Login to comment
183 Panel A: Inverted-membrane vesicles (200 µg) containing either wild-type YvcC (wt) or the K380R YvcC mutant (K380R) were added to the cuvette and after ~1 min incubation at 37 °C, 2 µM Hoechst 33342 was added. Login to comment
192 or C41(DE3) bacteria overexpressing either wild-type YvcC (wt) or the K380R YvcC mutant (K380R) were used, and the 7-amino- actinomycin D transport was monitored at 37 °C. After addition of 10 µM 7-aminoactinomycin D (1-2 min), 2 mM ATP or 2 mM AMPPNP was added where indicated by the arrow. Login to comment