ABCB1 p.Ser35Ala
| Predicted by SNAP2: | A: D (63%), C: D (66%), D: D (66%), E: D (75%), F: D (71%), G: N (87%), H: D (71%), I: D (71%), K: D (71%), L: D (75%), M: D (71%), N: N (72%), P: N (66%), Q: D (75%), R: D (75%), T: D (53%), V: D (71%), W: D (80%), Y: D (75%), |
| Predicted by PROVEAN: | A: N, C: N, D: N, E: N, F: D, G: N, H: N, I: D, K: N, L: D, M: N, N: N, P: N, Q: N, R: N, T: N, V: D, W: N, Y: D, |
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[hide] Biochemical characterization of domains in the mem... Biochemistry. 2005 Feb 22;44(7):2661-70. Kaur P, Rao DK, Gandlur SM
Biochemical characterization of domains in the membrane subunit DrrB that interact with the ABC subunit DrrA: identification of a conserved motif.
Biochemistry. 2005 Feb 22;44(7):2661-70., 2005-02-22 [PMID:15709779]
Abstract [show]
DrrA and DrrB proteins confer resistance to the commonly used anticancer agents daunorubicin and doxorubicin in the producer organism Streptomyces peucetius. The drrAB locus has previously been cloned in Escherichia coli, and the proteins have been found to be functional in this host. DrrA, a soluble protein, belongs to the ABC family of proteins. It forms a complex with the integral membrane protein DrrB. Previous studies suggest that the function and stability of DrrA and DrrB are biochemically coupled. Thus, DrrA binds ATP only when it is in a complex with DrrB in the membrane. Further, DrrB is completely degraded if DrrA is absent. In the present study, we have characterized domains in DrrB that may be directly involved in interaction with DrrA. Several single-cysteine substitutions in DrrB were made. Interaction between DrrA and DrrB was studied by using a cysteine to amine chemical cross-linker that specifically cross-links a free sulfhydryl group in one protein (DrrB) to an amine in another (DrrA). We show here that DrrA cross-links with both the N- and the C-terminal ends of the DrrB protein, implying that they may be involved in interaction. Furthermore, this study identifies a motif within the N-terminal cytoplasmic tail of DrrB, which is similar to a motif recently shown by crystal structure analysis in BtuC and previously shown by sequence analysis to be also present in exporters, including MDR1. We propose that the motif present in DrrB and other exporters is actually a modified version of the EAA motif, which was originally believed to be present only in the importers of the ABC family. The present work is the first report where domains of interaction in the membrane component of an ABC drug exporter have been biochemically characterized.
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No. Sentence Comment
253 These changes include S23A, G25A, E26D, and S35A in the N-terminal domain and C260A and A270S in the C-terminal domain.
X
ABCB1 p.Ser35Ala 15709779:253:44
status: NEW255 Since S35A mutation in the N-terminal domain and C260A and A270S in the C-terminal domain showed no change in the doxorubicin resistance phenotype as compared to the wild type, nonconservative mutations of these three residues, including S35I, C260E, and A270Y, were created.
X
ABCB1 p.Ser35Ala 15709779:255:6
status: NEW257 Protein expression analysis by SDS-PAGE showed that most mutations, except S23A, S35A, and A270Y show varying degrees of reduction in the levels of DrrA and DrrB.
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ABCB1 p.Ser35Ala 15709779:257:81
status: NEW