Slow-phase kinetics of nucleotide binding to the uncoupling protein from brown adipose tissue mitochondria.

The Journal of Biological Chemistry
S G HuangM Klingenberg

Abstract

The kinetics of nucleotide binding to the uncoupling protein (UCP) from brown adipose tissue mitochondria were studied with a filter binding method. Fast and slow phases of binding were observed, corresponding to the two-stage binding model based on equilibrium binding studies (Huang, S. G., and Klingenberg, M. (1996) Biochemistry 35, 7846-7854) (Reaction 1). [reaction: see text] Although this method determines total binding, only the slow phase can be resolved. The fast unresolved phase represents the formation of the initial loose UCP-nucleotide complex (UN; Kd approximately 2 microM), whereas the slow phase reflects the tight binding (U*N) associated with a conformational change induced by the bound nucleotide. Best fits of the binding data yielded, for the slow phase, k+1 values of 3.0 x 10(-3) s-1 for GTP, 4.8 x 10(-3) s-1 for ATP, 0.13 s-1 for GDP, and >0.7 s-1 for ADP and dissociation rate constants (k-1) of 0.10 x 10(-3) s-1 for GTP, 0.58 x 10(-3) s-1 for ATP, 8.8 x 10(-3) s-1 for GDP, and >0.3 s-1 for ADP at pH 6.7 and 4 degrees C. The rates were fairly pH- and temperature-dependent. The distribution constant Kc' (=k+1/k-1) between the tight and loose complexes ranged between 2 and 30, suggesting formation of 71-97% of...Continue Reading

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Citations

Mar 21, 2003·Archives of Biochemistry and Biophysics·Shu-Gui Huang
Jun 30, 2006·The International Journal of Biochemistry & Cell Biology·Petr JezekMartin Jabůrek
Apr 20, 2016·Biochimica Et Biophysica Acta·Frédéric BouillaudDaniel Ricquier
Mar 10, 2001·Biochimica Et Biophysica Acta·M Klingenberg, K S Echtay
Jun 20, 2006·Biochimica Et Biophysica Acta·Andrea DlaskováPetr Jezek
Feb 24, 2010·Biochimica Et Biophysica Acta·Martin Klingenberg
Oct 31, 2016·Biochimie·Martin Klingenberg
Jul 20, 2007·Magnetic Resonance in Chemistry : MRC·Patrick GrovesJesús Jiménez-Barbero
Jan 16, 1999·Biochimica Et Biophysica Acta·M Klingenberg, S G Huang

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