Biphasic Proton Transport Mechanism for Uncoupling Proteins

被引:7
作者
Ardalan, Afshan [1 ]
Sowlati-Hashjin, Shahin [2 ,3 ]
Oduwoye, Habib [1 ]
Uwumarenogie, Stephanie O. [1 ]
Karttunen, Mikko [2 ,3 ,4 ]
Smith, Matthew D. [5 ]
Jelokhani-Niaraki, Masoud [1 ]
机构
[1] Wilfrid Laurier Univ, Dept Chem & Biochem, Waterloo, ON N2L 3C5, Canada
[2] Univ Western Ontario, Dept Chem, London, ON N6A 3K7, Canada
[3] Univ Western Ontario, Ctr Adv Mat & Biomat Res, London, ON N6K 3K7, Canada
[4] Univ Western Ontario, Dept Phys & Astron, London, ON N6A 3K7, Canada
[5] Wilfrid Laurier Univ, Dept Biol, Waterloo, ON N2L 3C5, Canada
基金
加拿大自然科学与工程研究理事会; 加拿大创新基金会;
关键词
PARTICLE MESH EWALD; MOLECULAR-DYNAMICS; MITOCHONDRIAL CARRIERS; ALPHA-HEMOLYSIN; UCP2; EXPRESSION; BINDING; CONFORMATION; MUTAGENESIS; SIMULATIONS;
D O I
10.1021/acs.jpcb.1c04766
中图分类号
O64 [物理化学(理论化学)、化学物理学];
学科分类号
070304 ; 081704 ;
摘要
It has been suggested that uncoupling proteins (UCPs) transport protons via interconversion between two conformational states: one in the "cytoplasmic state" and the other in the "matrix state". Matrix and cytoplasmic salt-bridge networks are key controllers of these states. This study proposes a mechanism for proton transport in tetrameric UCP2, with focus on the role of the matrix network. Eleven mutants were prepared to disrupt (K -> Q or D -> N mutations) or alter (K -> D and D -> K mutations) the salt-bridges in the matrix network. Proteins were recombinantly expressed in Escherichia coli membrane, reconstituted in model lipid membranes, and their structures and functions were analyzed by gel electrophoresis, circular dichroism spectroscopy, fluorescence assays, as well as molecular dynamics simulations. It is shown that the UCP2 matrix network contains five salt-bridges (rather than the previously reported three), and the matrix network can regulate the proton transport by holding the protein's transmembrane helices in close proximity, limiting the movement of the activator fatty acid(s). A biphasic two-state molecular model is proposed for proton transport in tetrameric (a dimer of stable dimers) UCP2, in which all the monomers are functional, and monomers in each dimer are in the same transport mode. Purine nucleotide (e.g., ATP) can occlude the internal pore of the monomeric units of UCP tetramers via interacting with positive residues at or in the proximity of the matrix network (K38, K141, K239, R88, R185, and R279) and prevent switching between cytoplasmic and matrix states, thus inhibiting the proton transport. This study provides new insights into the mechanism of proton transport and regulation in UCPs.
引用
收藏
页码:9130 / 9144
页数:15
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