ABCC1 p.Cys7Ala
| Predicted by SNAP2: | A: D (63%), D: D (85%), E: D (80%), F: D (80%), G: D (75%), H: D (71%), I: D (75%), K: D (85%), L: D (75%), M: D (80%), N: D (75%), P: D (91%), Q: D (80%), R: D (80%), S: D (66%), T: D (75%), V: D (75%), W: D (85%), Y: D (75%), |
| Predicted by PROVEAN: | A: D, D: N, E: D, F: D, G: D, H: D, I: D, K: D, L: D, M: D, N: D, P: D, Q: D, R: D, S: D, T: D, V: D, W: D, Y: D, |
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[hide] Structural and functional consequences of mutating... J Biol Chem. 2002 Nov 15;277(46):44268-77. Epub 2002 Sep 13. Yang Y, Chen Q, Zhang JT
Structural and functional consequences of mutating cysteine residues in the amino terminus of human multidrug resistance-associated protein 1.
J Biol Chem. 2002 Nov 15;277(46):44268-77. Epub 2002 Sep 13., 2002-11-15 [PMID:12235150]
Abstract [show]
Multidrug resistance-associated protein 1 (MRP1) is a member of the ATP-binding cassette membrane transport superfamily and is responsible for multidrug resistance in cancer cells. Currently, there are nine known human MRPs. Distinct from many other members of the ATP-binding cassette superfamily, human MRP1 and four other MRPs have an additional membrane-spanning domain (MSD) with a putative extracellular amino terminus. The functional significance of this additional MSD (MSD1) is currently unknown. To understand the role of MSD1 in human MRP1 structure and function, we studied the amino-terminal 33 amino acids. We found that the amino terminus of human MRP1 has two cysteine residues (Cys(7) and Cys(32)) that are conserved among the five human MRPs that have MSD1. Mutation analyses of the two cysteines in human MRP1 revealed that the Cys(7) residue is critical for the MRP1-mediated drug resistance and leukotriene C(4) transport activity. On the other hand, mutation of Cys(32) reduced only moderately the MRP1 function. The effect of Cys(7) mutation on MRP1 activity appears to be due to the 5-7-fold decrease in the maximal transport rate V(max). We also found that mutation of Cys(7) changed the amino-terminal conformation of MRP1. This conformational change is likely responsible for the decrease in V(max) of LTC(4) transport mediated by the mutant MRP1. Based on these studies, we conclude that the amino terminus of human MRP1 is important and that the Cys(7) residue plays a critical role in maintaining the proper structure and function of human MRP1.
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No. Sentence Comment
49 Engineering Human MRP1 Constructs-Mutations of Cys7 and Cys32 to Ala were generated using the TransformerTM site-directed mutagenesis kit (Clontech, Palo Alto, CA).
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ABCC1 p.Cys7Ala 12235150:49:47
status: NEW[hide] Functional and structural consequences of cysteine... Biochemistry. 2003 May 13;42(18):5214-24. Leslie EM, Letourneau IJ, Deeley RG, Cole SP
Functional and structural consequences of cysteine substitutions in the NH2 proximal region of the human multidrug resistance protein 1 (MRP1/ABCC1).
Biochemistry. 2003 May 13;42(18):5214-24., 2003-05-13 [PMID:12731862]
Abstract [show]
The 190 kDa multidrug resistance protein 1 (MRP1; ABCC1) is comprised of three membrane spanning domains (MSDs) and two nucleotide binding domains (NBDs) configured MSD1-MSD2-NBD1-MSD3-NBD2. MRP1 overexpression in tumor cells results in an ATP-dependent efflux of many oncolytic agents and arsenic and antimony oxyanions. MRP1 also transports GSSG and GSH as well as conjugated organic anions, including leukotriene C(4) and 17beta-estradiol 17-(beta-D-glucuronide) and certain xenobiotics in association with GSH. Previous studies have shown that portions of MSD1 and the cytoplasmic loop (CL3) connecting it to MSD2 are important for MRP1 transport function. In the present study, Cys residues at positions 43, 49, 85, 148, and 190 in MSD1 and positions 208 and 265 in CL3 were mutated to Ala and Ser, and the effects on protein expression, plasma membrane localization, trypsin sensitivity, organic anion transport, and drug resistance properties were investigated. Confocal microscopy showed that 11 of 14 mutants displayed significant levels of nonplasma membrane-associated MRP1. Most mutant proteins were also more resistant to trypsin proteolysis than wild-type MRP1. All Cys mutants transported organic anions (0.5-1.5-fold wild-type MRP1 activity), and cells expressing Ser-substituted but not Ala-substituted Cys43 and Cys265 MRP1 mutants exhibited a 2.5-fold decrease and a 3-fold increase in arsenite resistance, respectively; Cys43Ser MRP1 also conferred lower levels of vincristine resistance. These results indicate that certain Cys residues in the NH(2) proximal region of MRP1 can be important for its structure and selected transport activities.
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No. Sentence Comment
222 Yang et al. (26) recently reported that when Cys7 and Cys32 in the extracellular NH2 terminus of MRP1 were replaced with Ala, the Cys7Ala but not the Cys32Ala mutant showed significant structural and functional changes.
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ABCC1 p.Cys7Ala 12731862:222:130
status: NEW224 However, in contrast to our findings with the seven Cys mutants examined here, LTC4 transport by Cys7Ala-MRP1 was reduced by more than 90%, providing indirect evidence that the structure of MRP1 was more disrupted in this mutant.
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ABCC1 p.Cys7Ala 12731862:224:97
status: NEW225 On the other hand, the apparent changes in Cys7Ala-MRP1 conformation resulted in little or no impairment of plasma membrane localization unlike several of the Cys-substituted mutants investigated here (26).
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ABCC1 p.Cys7Ala 12731862:225:43
status: NEW[hide] Mutation of proline residues in the NH(2)-terminal... Biochim Biophys Acta. 2003 Sep 2;1615(1-2):103-14. Ito K, Weigl KE, Deeley RG, Cole SP
Mutation of proline residues in the NH(2)-terminal region of the multidrug resistance protein, MRP1 (ABCC1): effects on protein expression, membrane localization, and transport function.
Biochim Biophys Acta. 2003 Sep 2;1615(1-2):103-14., [PMID:12948592]
Abstract [show]
The Multidrug Resistance Protein, MRP1 (ABCC1) confers drug resistance and transports organic anions such as leukotriene C(4) (LTC(4)) and 17beta-estradiol 17-(beta-D-glucuronide) (E(2)17betaG). Previous studies showed that portions of the first membrane spanning domain (MSD1) and the cytoplasmic loop (CL3) connecting it to MSD2 are important for MRP1 transport function. We have replaced 12 prolines in MSD1 and CL3 with alanine and determined the effects of these substitutions on MRP1 expression and transport activity. All singly substituted MRP1-Pro mutants could be expressed in HeLa cells, except MRP1-P104A. The expressed mutants also transported LTC(4) and E(2)17betaG, and their K(m) (LTC(4)) values were similar to wild-type MRP1. Expression of the double mutant MRP1-P42/51A was reduced by >80% although it localized to the plasma membrane and transported organic anions. MRP1 expression was also reduced when the first transmembrane helix (amino acids 37-54) was deleted. In contrast, the phenotypes of the multiply substituted CL3 mutants MRP1-P196/205/207/209A and MRP1-P235/255A were comparable to wild-type MRP1. However, Pro(255)-substituted MRP1 mutants showed reduced immunoreactivity with a monoclonal antibody (MAb) whose epitope is located in CL3. We conclude that certain prolines in MSD1 and CL3 play a role in the expression and structure of MRP1.
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No. Sentence Comment
222 Consistent with this idea are the recent findings of Yang et al. [36] which showed that replacement of Cys7 with Ala caused an almost complete loss of LTC4 transport activity and a change in conformation of the NH2 terminus although expression levels and plasma membrane localization of this mutant MRP1 were not diminished compared to wild-type MRP1.
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ABCC1 p.Cys7Ala 12948592:222:103
status: NEW221 Consistent with this idea are the recent findings of Yang et al. [36] which showed that replacement of Cys7 with Ala caused an almost complete loss of LTC4 transport activity and a change in conformation of the NH2 terminus although expression levels and plasma membrane localization of this mutant MRP1 were not diminished compared to wild-type MRP1.
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ABCC1 p.Cys7Ala 12948592:221:103
status: NEW