Mutations of Charged Amino Acids in or near the Transmembrane Helices of the Second Membrane Spanning Domain Differentially Affect the Substrate Specificity and Transport Activity of the Multidrug Resistance Protein MRP1 (ABCC1)

2004 ◽  
Vol 65 (6) ◽  
pp. 1375-1385 ◽  
Author(s):  
Anass Haimeur ◽  
Gwenaëlle Conseil ◽  
Roger G. Deeley ◽  
Susan P.C. Cole
Keyword(s):  
Amino Acids ◽  
Multidrug Resistance ◽  
Substrate Specificity ◽  
Multidrug Resistance Protein ◽  
Transmembrane Helices ◽  
Transport Activity ◽  
Charged Amino Acids ◽  
Resistance Protein ◽  
Membrane Spanning

scholarly journals Multiple Membrane-associated Tryptophan Residues Contribute to the Transport Activity and Substrate Specificity of the Human Multidrug Resistance Protein, MRP1

2002 ◽  
Vol 277 (51) ◽  
pp. 49495-49503 ◽  
Author(s):  
Koji Koike ◽  
Curtis J. Oleschuk ◽  
Anass Haimeur ◽  
Sharon L. Olsen ◽  
Roger G. Deeley ◽  
...  
Keyword(s):  
Multidrug Resistance ◽  
Substrate Specificity ◽  
Organic Anion ◽  
Selective Reduction ◽  
Multidrug Resistance Protein ◽  
Transport Activity ◽  
Binding Domains ◽  
17Β Estradiol ◽  
Resistance Protein ◽  
Estradiol 17Β

The multidrug resistance protein, MRP1, is a clinically important ATP-binding cassette transporter in which the three membrane-spanning domains (MSDs), which contain up to 17 transmembrane (TM) helices, and two nucleotide binding domains (NBDs) are configured MSD1-MSD2-NBD1-MSD3-NBD2. In tumor cells, MRP1 confers resistance to a broad spectrum of drugs, but in normal cells, it functions as a primary active transporter of organic anions such as leukotriene C4and 17β-estradiol 17β-(d-glucuronide). We have previously shown that mutation of TM17-Trp1246eliminates 17β-estradiol 17β-(d-glucuronide) transport and drug resistance conferred by MRP1 while leaving leukotriene C4transport intact. By mutating the 11 remaining Trp residues that are in predicted TM segments of MRP1, we have now determined that five of them are also major determinants of MRP1 function. Ala substitution of three of these residues, Trp445(TM8), Trp553(TM10), and Trp1198(TM16), eliminated or substantially reduced transport levels of five organic anion substrates of MRP1. In contrast, Ala substitutions of Trp361(TM7) and Trp459(TM9) caused a more moderate and substrate-selective reduction in MRP1 function. More conservative substitutions (Tyr and Phe) of the Trp445, Trp553, and Trp1198mutants resulted in substrate selective retention of transport in some cases (Trp445and Trp1198) but not others (Trp553). Our findings suggest that the bulky polar aromatic indole side chain of each of these five Trp residues contributes significantly to the transport activity and substrate specificity of MRP1.

scholarly journals The First Cytoplasmic Loop in the Core Structure of the ABCC1 (Multidrug Resistance Protein 1; MRP1) Transporter Contains Multiple Amino Acids Essential for Its Expression

2021 ◽  
Vol 22 (18) ◽  
pp. 9710
Author(s):  
Gwenaëlle Conseil ◽  
Susan P. C. Cole
Keyword(s):  
Amino Acids ◽  
Multidrug Resistance ◽  
Core Structure ◽  
Multidrug Resistance Protein ◽  
Cytoplasmic Loop ◽  
Multidrug Resistance Protein 1 ◽  
Resistance Protein ◽  
Membrane Spanning ◽  
Α Helix ◽  
Membrane Spanning Domains

ABCC1 (human multidrug resistance protein 1 (hMRP1)) is an ATP-binding cassette transporter which effluxes xeno- and endobiotic organic anions and confers multidrug resistance through active drug efflux. The 17 transmembrane α-helices of hMRP1 are distributed among three membrane spanning domains (MSD0, 1, 2) with MSD1,2 each followed by a nucleotide binding domain to form the 4-domain core structure. Eight conserved residues in the first cytoplasmic loop (CL4) of MSD1 in the descending α-helix (Gly392, Tyr404, Arg405), the perpendicular coupling helix (Asn412, Arg415, Lys416), and the ascending α-helix (Glu422, Phe434) were targeted for mutagenesis. Mutants with both alanine and same charge substitutions of the coupling helix residues were expressed in HEK cells at wild-type hMRP1 levels and their transport activity was only moderately compromised. In contrast, mutants of the flanking amino acids (G392I, Y404A, R405A/K, E422A/D, and F434Y) were very poorly expressed although Y404F, E422D, and F434A were readily expressed and transport competent. Modeling analyses indicated that Glu422 and Arg615 could form an ion pair that might stabilize transporter expression. However, this was not supported by exchange mutations E422R/R615E which failed to improve hMRP1 levels. Additional structures accompanied by rigorous biochemical validations are needed to better understand the bonding interactions crucial for stable hMRP1 expression.

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