Abstract
Structural analyses of bacterial ATP-binding-cassette transporters revealed that the glutamine residue in Q-loop plays roles in interacting with: 1) a metal cofactor to participate in ATP binding; 2) a putative catalytic water molecule to participate in ATP hydrolysis; 3) other residues to transmit the conformational changes between nucleotide-binding-domains and transmembrane-domains, in ATP-dependent solute transport. We have mutated the glutamines at 713 and 1375 to asparagine, methionine or leucine to determine the functional roles of these residues in Q-loops of MRP1. All these single mutants significantly decreased Mg • ATP binding and increased the Km (Mg • ATP) and Vmax values in Mg • ATP-dependent leukotriene-C4 transport. However, the Vmax values of the double mutants Q713N/Q1375N, Q713M/Q1375M and Q713L/Q1375L were lower than that of wtMRP1, implying that the double mutants cannot efficiently bind Mg • ATP. Interestingly, MRP1 has higher affinity for Mn • ATP than for Mg • ATP and the Mn • ATP-dependent leukotriene-C4 transport activities of Q713N/Q1375N and Q713M/Q1375M are significantly higher than that of wtMRP1. All these results suggest that: 1) the glutamine residues in Q-loops contribute to ATP-binding via interaction with a metal cofactor; 2) it is most unlikely that these glutamine residues would play crucial roles in ATP hydrolysis and in transmitting the conformational changes between nucleotide-binding-domains and transmembrane-domains.
Original language | English (US) |
---|---|
Pages (from-to) | 1790-1796 |
Number of pages | 7 |
Journal | Biochimica et Biophysica Acta - Biomembranes |
Volume | 1808 |
Issue number | 7 |
DOIs | |
State | Published - Jul 2011 |
Keywords
- ATP binding/hydrolysis
- ATP dependent
- Glutamine residue in Q-loop
- K and V values
- LTC4 transport
- MRP1
- Metal cofactor
ASJC Scopus subject areas
- Biophysics
- Biochemistry
- Cell Biology