Binding of ATP to uncoupling protein of brown fat mitochondria as studied by means of spin‐labeled ATP derivatives

FEBS Letters

Jakobs, P.²  Braun, A.²  Trommer, W.E.²  Jezek, P.¹ 

[1] Institute of Physiology, Czechoslovak Academy of Sciences, Videnska 1083, 14220 Prague, Czechoslovakia;Fachbereich Chemie, Universität Kaiserslautern, Erwin-Schrödinger Straße, D-6750 Kaiserslautern, Germany

关键词: Uncoupling protein;    Chemical modification;    Brown fat mitochondria;    ATP binding site;    Spin-labeled ATP;    UCP;    uncoupling protein;    C8-SL;    preceding ATP or AMP refers to their 8-(2;    2;    6;    6-tetramethylpiperidin-4-yl-1-oxyl)amino derivatives;    N6-SL;    preceding ATP or AMP refers to their N 6-(2;    2;    6;    6-tetramethylpiperidin-4-yl-1-oxyl) derivatives;    C2′;    3′-SL preceding ATP refers to the equilibrium mixture of its C2′- and C3′-(2;    2;    5;    5-tetramethyl-3-pyrroline-1-oxyl-3-carboxylic acid esters);    UCP;    uncoupling protein from brown adipose tissue;    NEM;    N-ethylmaleimide;    DABS;    diazobenzene-4-sulfonic acid;    HEPES;    N-(2-h;    droxyethyl) 1-piperazineethanesulfonic acid;    EGTA;    ethylenebis (oxyethylenenitrilo)tetraacetate;    MES;    2-morpholinoethanesulfonic acid;   

DOI  :  10.1016/0014-5793(91)80683-T

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

ATP derivatives spin-labeled (SL) at C8, N⁶, C2′ or C3′ were employed in binding studies with the uncoupling protein of brown fat mitochondria. Substitution of the ribose strongly impaired binding, whereas labeling of the adenine moiety allowed for tight and functional complex formation. Detailed binding studies with C8-SL-ATP confirmed the known pH and Mg²⁺ dependence with a stoichiometry of one C8-SL-ATP bound per 66 kDa dimer. Corresponding studies of the uncoupling protein after modification with N-ethylmaleimide or diazobenzene-4-sulfonic acid revealed distinct differences in their effects on nucleotide binding and gating.

【 授权许可】

Unknown