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PMID:24196960

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Citation

Hoang, T, Smith, MD and Jelokhani-Niaraki, M' (2013) Expression, folding and proton transport activity of human uncoupling protein-1(UCP1) in lipid membranes: evidence for associated functional forms. J. Biol. Chem. '

Abstract

Uncoupling protein-1 (UCP1) is abundantly expressed in the mitochondrial inner membrane of brown adipose tissues (BAT) and has an important role in heat generation, mediated by its proton transport function. The structure and function of UCP1 are not fully understood, partially due to the difficulty in obtaining native-like folded proteins in vitro. In this study, using the auto-induction method, we have successfully expressed UCP1 in E. coli membranes in high yield. Overexpressed UCP1 in bacterial membranes was extracted using mild detergents and reconstituted into phospholipid bilayers for biochemical studies. UCP1 was folded in octyl glucoside, as indicated by its high helical content and binding to ATP, a known UCP1 proton transport inhibitor. Reconstituted UCP1 in phospholipid vesicles also exhibited highly helical structures and proton transport that is activated by fatty acids and inhibited by purine nucleotides. Self-associated functional forms of UCP1 in lipid membranes were observed for the first time. The self-assembly of UCP1 into tetramers was unambiguously characterized by circular dichroism and fluorescence spectroscopy, analytical ultracentrifugation and semi-native gel electrophoresis. In addition, the mitochondrial lipid cardiolipin stabilized the structure of associated UCP1 and enhanced the proton transport activity of the protein. The existence of the functional oligomeric states of UCP1 in the lipid membranes has important implications for understanding the structure and proton transport mechanism of this protein in BAT, as well as structure-function relationships of other mammalian UCPs in other tissues.

Links

PubMed Online version:10.1074/jbc.M113.509935

Keywords


Significance

Annotations

Gene product Qualifier GO Term Evidence Code with/from Aspect Extension Notes Status

HUMAN:UCP1

GO:0017077: oxidative phosphorylation uncoupler activity

ECO:0000314:

F

Figure 2: Measures the expression of UCP1 and NADH activity in the bacterial membranes and supernatant.

complete
CACAO 9190

HUMAN:UCP1

enables

GO:0022857: transmembrane transporter activity

ECO:0000314: direct assay evidence used in manual assertion

F

Seeded From UniProt

complete

HUMAN:UCP1

involved_in

GO:1902600: proton transmembrane transport

ECO:0000314: direct assay evidence used in manual assertion

P

Seeded From UniProt

complete

See also

References

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