ABCMdb

PMID: 15182191

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 [PubMed]

Sentences

No. Mutations Sentence Comment

121 ABCB1 p.Lys380Arg ABCB1 p.Lys380Arg 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 122 ABCB1 p.Lys380Ala A similar result was also obtained with the K380A mutant, which is totally devoid of ATPase activity (not shown). Login to comment 132 ABCB1 p.Lys380Arg 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 173 ABCB1 p.Lys380Ala Furthermore, no ATPase activity was detected when purified E504A, E504Q [corresponding to the catalytic base of ABC transporters (47)], or K380A YvcC mutant was used, ruling out the possibility that the ATPase activity seen with the wild-type YvcC might be due to a contaminant. Login to comment 183 ABCB1 p.Lys380Arg ABCB1 p.Lys380Arg 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 ABCB1 p.Lys380Arg ABCB1 p.Lys380Arg 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