Guanine nucleotide transport by atractyloside-sensitive and -insensitive carriers in isolated heart mitochondria.
In previous work (McKee EE, Bentley AT, Smith RM Jr, and Ciaccio CE, Biochem Biophys Res Commun 257: 466-472, 1999), the transport of guanine nucleotides into the matrix of intact isolated heart mitochondria was demonstrated. In this study, the time course and mechanisms of guanine nucleotide transport are characterized. Two distinct mechanisms of transport were found to be capable of moving guanine nucleotides across the inner membrane. The first carrier was saturable, displayed temperature dependence, preferred GDP to GTP, and did not transport GMP or IMP. When incubated in the absence of exogenous ATP, this carrier had a V(max) of 946 +/- 53 pmol. mg(-1). min(-1) with a K(m) of 2.9 +/- 0.3 mM for GDP. However, transport of GTP and GDP on this carrier was completely inhibited by physiological concentrations of ATP, suggesting that this carrier was not involved with guanine nucleotide transport in vivo. Because transport on this carrier was also inhibited by atractyloside, this carrier was consistent with the well-characterized ATP/ADP translocase. The second mechanism of guanine nucleotide uptake was insensitive to atractyloside, displayed temperature dependence, and was capable of transporting GMP, GDP, and GTP at approximately equal rates but did not transport IMP, guanine, or guanosine. GTP transport via this mechanism was slow, with a V(max) of 48.7 +/- 1.4 pmol. mg(-1). min(-1) and a K(m) = 4.4 +/- 0.4 mM. However, because the requirement for guanine nucleotide transport is low in nondividing tissues such as the heart, this transport process is nevertheless sufficient to account for the matrix uptake of guanine nucleotides and may represent the physiological mechanism of transport.
McKee
E E
EE
South Bend Center for Medical Education, Indiana University School of Medicine, University of Notre Dame, Notre Dame, Indiana 46556, USA. mcKee.6@nd.edu
Bentley
A T
AT
Smith
R M
RM
Kraas
J R
JR
Ciaccio
C E
CE
D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D001278:Atractyloside / Q000494:pharmacology*; D001692:Biological Transport / Q000187:drug effects / Q000502:physiology; D004305:Dose-Response Relationship, Drug; D004734:Energy Metabolism / Q000502:physiology; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005033:Ethylmaleimide / Q000494:pharmacology; D006150:Guanine Nucleotides / Q000493:pharmacokinetics*; D006153:Guanosine Diphosphate / Q000493:pharmacokinetics; D006157:Guanosine Monophosphate / Q000493:pharmacokinetics; D006160:Guanosine Triphosphate / Q000493:pharmacokinetics; D006902:Hydroxymercuribenzoates / Q000494:pharmacology; D007700:Kinetics; D008928:Mitochondria / Q000378:metabolism*; D009206:Myocardium / Q000378:metabolism*; D011480:Protease Inhibitors / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D014316:Tritium
D004791:Enzyme Inhibitors; D006150:Guanine Nucleotides; D006902:Hydroxymercuribenzoates; D011480:Protease Inhibitors; D014316:Tritium; C030601:4-hydroxymercuribenzoate; D006153:Guanosine Diphosphate; D001278:Atractyloside; D006157:Guanosine Monophosphate; D006160:Guanosine Triphosphate; D000255:Adenosine Triphosphate; D005033:Ethylmaleimide