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Characterization of Nucleotide Transport into Rat Brain Synaptic Vesicles
Author(s) -
Gualix Javier,
Pintor Jesús,
MirasPortugal Maria Teresa
Publication year - 1999
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1046/j.1471-4159.1999.0731098.x
Subject(s) - nucleotide , allosteric regulation , adenosine triphosphate , saturation vapor curve , adenosine , chemistry , synaptic vesicle , adenine nucleotide , vesicle , biochemistry , stereochemistry , adenosine diphosphate , biophysics , enzyme , biology , membrane , platelet , platelet aggregation , immunology , gene
: ATP transport to synaptic vesicles from rat brain has been studied using the fluorescent substrate analogue 1, N 6 ‐ethenoadenosine 5′‐triphosphate (ε‐ATP), The increase in intravesicular concentration was time dependent for the first 30 min, ε‐ATP being the most abundant nucleotide. The complexity of the saturation curve indicates the existence of kinetic and allosteric cooperativity in the nucleotide transport, which exhibits various affinity states with K 0.5 values of 0.39 ± 0.06 and 3.8 ± 0.1 m M with ε‐ATP as substrate. The V max values obtained were 13.5 ± 1.4 pmol · min ‐1 · mg of protein ‐1 for the first curve and 28.3 ± 1.6 pmol · min ‐1 · mg of protein ‐1 considering both components. This kinetic behavior can be explained on the basis of a mnemonic model. The nonhydrolyzable adenine nucleotide analogues adenosine 5′‐ O ‐3‐(thiotriphosphate), adenosine 5′‐(β,γ‐imino)triphosphate and the diadenosine polyphosphates P 1 , P 3 ‐di(adenosine)triphosphate, P 1 , P 4 ‐di(adenosine)tetraphosphate, and P 1 , P 5 ‐di(adenosine)pentaphosphae the nucleotide transport. The mitochondrial ATP/ADP exchange inhibitor atractyloside, N ‐ethylmaleimide, and polysulfonic aromatic compounds such as Evans blue and 4,4′‐diisothiocyanostilbene‐2,2′‐disulfonic acid also inhibit ε‐ATP vesicular transport.