ABCMdb

ABCB1 p.Glu1013Gly

Predicted by SNAP2:	A: N (53%), C: D (53%), D: N (97%), F: D (66%), G: D (66%), H: D (53%), I: D (66%), K: N (57%), L: D (63%), M: D (63%), N: N (87%), P: D (80%), Q: N (53%), R: N (61%), S: N (66%), T: D (59%), V: D (63%), W: D (71%), Y: D (66%),
Predicted by PROVEAN:	A: D, C: D, D: N, F: D, G: D, H: D, I: D, K: D, L: D, M: D, N: D, P: D, Q: N, R: D, S: D, T: D, V: D, W: D, Y: D,

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Publications
[hide] Generating symmetry in the asymmetric ATP-binding ... J Biol Chem. 2014 May 30;289(22):15272-9. doi: 10.1074/jbc.M114.553065. Epub 2014 Apr 14. Gupta RP, Kueppers P, Hanekop N, Schmitt L
Generating symmetry in the asymmetric ATP-binding cassette (ABC) transporter Pdr5 from Saccharomyces cerevisiae.
J Biol Chem. 2014 May 30;289(22):15272-9. doi: 10.1074/jbc.M114.553065. Epub 2014 Apr 14., [PMID:24733388]

Abstract [show]
Pdr5 is a plasma membrane-bound ABC transporter from Saccharomyces cerevisiae and is involved in the phenomenon of resistance against xenobiotics, which are clinically relevant in bacteria, fungi, and humans. Many fungal ABC transporters such as Pdr5 display an inherent asymmetry in their nucleotide-binding sites (NBS) unlike most of their human counterparts. This degeneracy of the NBSs is very intriguing and needs explanation in terms of structural and functional relevance. In this study, we mutated nonconsensus amino acid residues in the NBSs to its consensus counterpart and studied its effect on the function of the protein and effect on yeast cells. The completely "regenerated" Pdr5 protein was severely impaired in its function of ATP hydrolysis and of rhodamine 6G transport. Moreover, we observe alternative compensatory mechanisms to counteract drug toxicity in some of the mutants. In essence, we describe here the first attempts to restore complete symmetry in an asymmetric ABC transporter and to study its effects, which might be relevant to the entire class of asymmetric ABC transporters.

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No. Sentence Comment
71 Mutation Direction Sequence, 5d15; to 3d15; P195S S GCTAGTCGTTTTAGGTAGATCAGGCTCTGGCAAAAC AS GTTTTGCCAGAGCCTGATCTACCTAAAACGACTAGC C199K S CGTTTTAGGTAGACCAGGCTCTGGCAAAACTACTTTATTAAAATCCATCTCTTC AS GAAGAGATGGATTTTAATAAAGTAGTTTTGCCAGAGC CTGGTCTACCTAAAACG N334E S GATCCAAATTTCAATGCTGGGATGAAGCTACAAGGG GTTTGGATTC AS GAATCCAAACCCCTTGTAGCTTCATCCCAGCATTGAA ATTTGGATC Y367H S CTGCCACAGTGGCCATCCATCAATGTTCTCAAGATG AS CATCTTGAGAACATTGATGGATGGCCACTGTGGCAG N1011S S GTTGTTGGTGTTGCTGGTGAAGGTTTATCTGTTGAAC AAAGAAAAAGATTAACCATTG AS CAATGGTTAATCTTTTTCTTTGTTCAACAGATAAACCT TCACCAGCAACACCAACAAC V1012G S GTTGCTGGTGAAGGTTTAAACGGTGAACAAAGAAAA AGATTAACC AS GGTTAATCTTTTTCTTTGTTCACCGTTTAAACCTTCAC CAGCAAC E1013G S GTTGCTGGTGAAGGTTTAAACGTTGGTCAAAGAAAAA GATTAACCATTGGTG AS CACCAATGGTTAATCTTTTTCTTTGACCAACGTTTAAA CCTTCACCAGCAAC Role of Degenerate Nucleotide-binding Site in Pdr5 MAY 30, 2014ߦVOLUME 289ߦNUMBER 22 JOURNAL OF BIOLOGICAL CHEMISTRY 15273 at SEMMELWEIS UNIV OF MEDICINE on December 12, 40 mM H2SO4. Login to comment
86 In the following, restoration of the consensus sequence in the Walker A motif (C199K) located in NBD1 and forming NBS1 is called the "Walker A" mutant; restoration of the consensus sequence in the Walker B motif (N334E; NBD1) in NBS1 is called the "Walker B" mutant; restoration of the consensus sequence of the H-loop (Y367H, NBD1) in NBS1 is called the "H-loop" mutant, and restoration of the consensus sequence of the C-loop (N1011S/V1012G/ E1013G, NBD2) in NBS1 is called the "C-loop" mutant. Login to comment
112 TABLE 2 List of mutations generated and used in this study Mutation Abbreviation Alteration Motif Domain P195S Pro in Walker A Walker A NBD1 C199K Walker A Cys in Walker A Walker A NBD1 N334E Walker B Asn in Walker B Walker B NBD1 Y367H H-loop Tyr in H-loop H-loop NBD1 N1011S, V1012G, E1013G C-loop Asn, Val, and Glu in C-loop C-loop NBD2 Y367H, N1011S, V1012G, E1013G HC-loop Tyr in H-loop H-loop NBD1/2 Asn, Val, and Glu in C-loop C-loop C199K AHC Cys in Walker A Walker A NBD1/2 Y367H Tyr in H-loop H-loop N1011S, V1012G, E1013G Asn, Val, and Glu in C-loop C-loop P195S ABHC Pro in Walker A Walker A NBD1/2 C199K Cys in Walker A Walker B N334E Asn in Walker B H-loop Y367H Tyr in H-loop C-loop N1011S, V1012G, E1013G Asn, Val, and Glu in C-loop Role of Degenerate Nucleotide-binding Site in Pdr5 MAY 30, 2014ߦVOLUME 289ߦNUMBER 22 JOURNAL OF BIOLOGICAL CHEMISTRY 15275 at SEMMELWEIS UNIV OF MEDICINE on December 12, that the apparent affinity of these mutants toward ATP has not drastically changed (Table 3). Login to comment