Natural and azido fatty acids inhibit phosphate transport and activate fatty acid anion uniport mediated by the mitochondrial phosphate carrier.
The electroneutral P(i) uptake via the phosphate carrier (PIC) in rat liver and heart mitochondria is inhibited by fatty acids (FAs), by 12-(4-azido-2-nitrophenylamino)dodecanoic acid (AzDA) and heptylbenzoic acid ( approximately 1 microm doses) and by lauric, palmitic, or 12-azidododecanoic acids ( approximately 0.1 mm doses). In turn, reconstituted E. coli-expressed yeast PIC mediated anionic FA uniport with a similar pattern leading to FA cycling and H(+) uniport. The kinetics of P(i)/P(i) exchange on recombinant PIC in the presence of AzDA better corresponded to a competitive inhibition mechanism. Methanephosphonate was identified as a new PIC substrate. Decanephosphonate, butanephosphonate, 4-nitrophenylphosphate, and other P(i) analogs were not translocated and did not inhibit P(i) transport. However, methylenediphosphonate and iminodi(methylenephosphonate) inhibited both electroneutral P(i) uptake and FA cycling via PIC. AzDA analog 16-(4-azido-2-nitrophenylamino)-[(3)H(4)]-hexadecanoic acid ((3)H-AzHA) bound upon photoactivation to several mitochondrial proteins, including the 30- and 34-kDa bands. The latter was ascribed to PIC due to its specific elution pattern on Blue Sepharose and Affi-Gel. (3)H-AzHA photolabeling of recombinant PIC was prevented by methanephosphonate and diphosphonates and after premodification with 4-azido-2-nitrophenylphosphate. Hence, the demonstrated PIC interaction with monovalent long-chain FA anions, but with divalent phosphonates of short chain only, indicates a pattern distinct from that valid for the mitochondrial uncoupling protein-1.
Engstová
H
H
Institute of Physiology, Department of Membrane Transport Biophysics, Academy of Sciences of the Czech Republic, Prague CZ 14220, Czech Republic.
Zácková
M
M
Růzicka
M
M
Meinhardt
A
A
Hanus
J
J
Krämer
R
R
Jezek
P
P
D000345:Affinity Labels / Q000494:pharmacology; D000818:Animals; D001693:Biological Transport, Active; D002352:Carrier Proteins / Q000037:antagonists & inhibitors* / Q000378:metabolism*; D004164:Diphosphonates / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D005227:Fatty Acids / Q000378:metabolism / Q000494:pharmacology*; D017136:Ion Transport; D007700:Kinetics; D007850:Lauric Acids / Q000494:pharmacology*; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism; D010169:Palmitic Acids / Q000494:pharmacology; D028044:Phosphate-Binding Proteins; D010710:Phosphates / Q000378:metabolism*; D051381:Rats; D017208:Rats, Wistar
C098506:12-(4-azido-2-nitrophenylamino)dodecanoic acid; C098967:16-(4-azido-2-nitrophenylamino)hexadecanoic acid; D000345:Affinity Labels; D002352:Carrier Proteins; D004164:Diphosphonates; D005227:Fatty Acids; D007850:Lauric Acids; D010169:Palmitic Acids; D028044:Phosphate-Binding Proteins; D010710:Phosphates