Binding-protein-dependent transport systems in bacteria are members of the ATP-binding cassette (ABC) superfamily. ABC transporters secrete virulence factors in bacteria and contribute to the drug resistance of several pathogens. Genetic defects in human homologs are responsible for over a dozen serious human diseases including cystic fibrosis, adrenoleukodystrophy and cholestasis. The maltose transport system of E. coli is well characterized and is therefore an ideal model for the study of ABC transporters. A periplasmic MBP directs maltose to a membrane-associated transport complex containing two transrnembrane-spanning proteins, MalF and MalG, and two copies of a peripheral ATP-binding-protein, MaIK. ATP hydrolysis by the multisubunit complex drives maltose transport. The applicant's long-term objective is to understand how the structural and mechanistic features of the maltose transport system result in active transport. As this mechanism is complex, the approach is to stabilize intermediates in the pathway and characterize them using biochemical and biophysical approaches. In this project, the investigators will use site-directed spin labeling and EPR, along with other complementary techniques, to define the movements that occur in moving from one intermediate to the next. The purified preparations will also be used in collaborative efforts to obtain high-resolution structures of the multi-subunit complex in different conformational states.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
2R01GM049261-10
Application #
6611472
Study Section
Microbial Physiology and Genetics Subcommittee 2 (MBC)
Program Officer
Chin, Jean
Project Start
1993-05-01
Project End
2007-04-30
Budget Start
2003-05-01
Budget End
2004-04-30
Support Year
10
Fiscal Year
2003
Total Cost
$317,768
Indirect Cost
Name
Baylor College of Medicine
Department
Microbiology/Immun/Virology
Type
Schools of Medicine
DUNS #
051113330
City
Houston
State
TX
Country
United States
Zip Code
77030
Alvarez, Frances Joan D; Orelle, C├ędric; Huang, Yan et al. (2015) Full engagement of liganded maltose-binding protein stabilizes a semi-open ATP-binding cassette dimer in the maltose transporter. Mol Microbiol 98:878-94
Cui, Jinming; Qasim, Sabiha; Davidson, Amy L (2010) Uncoupling substrate transport from ATP hydrolysis in the Escherichia coli maltose transporter. J Biol Chem 285:39986-93
Alvarez, Frances Joan D; Orelle, Cedric; Davidson, Amy L (2010) Functional reconstitution of an ABC transporter in nanodiscs for use in electron paramagnetic resonance spectroscopy. J Am Chem Soc 132:9513-5
Orelle, Cedric; Alvarez, Frances Joan D; Oldham, Michael L et al. (2010) Dynamics of alpha-helical subdomain rotation in the intact maltose ATP-binding cassette transporter. Proc Natl Acad Sci U S A 107:20293-8
Davidson, Amy L; Alvarez, Frances Joan D (2010) Binding protein-dependent uptake of maltose into cells via an ATP-binding cassette transporter. EcoSal Plus 2010:
Khare, Dheeraj; Oldham, Michael L; Orelle, Cedric et al. (2009) Alternating access in maltose transporter mediated by rigid-body rotations. Mol Cell 33:528-36
Oldham, Michael L; Davidson, Amy L; Chen, Jue (2008) Structural insights into ABC transporter mechanism. Curr Opin Struct Biol 18:726-33
Lu, Gang; Westbrooks, James M; Davidson, Amy L et al. (2005) ATP hydrolysis is required to reset the ATP-binding cassette dimer into the resting-state conformation. Proc Natl Acad Sci U S A 102:17969-74
Davidson, Amy L; Chen, Jue (2004) ATP-binding cassette transporters in bacteria. Annu Rev Biochem 73:241-68
Chen, Jue; Lu, Gang; Lin, Jeffrey et al. (2003) A tweezers-like motion of the ATP-binding cassette dimer in an ABC transport cycle. Mol Cell 12:651-61