Purification and characterization of phosphate carrier from potato mitochondria

The phosphate carrier of the inner mitochondrial membrane has been purified from potato tuber in a nearly homogenous form and characterized after reco nstitution into liposomes. The mitochondrial membrane was solubilised by Tr iton X-114 in the presence of cardiolipin and the soluble fraction was chro matographied on hydroxylapatite. The flow-through fractions were collected and analyzed by SDS-gel electrophoresis. In some fractions, a single protei n band of M-r = 35 kDa was found. This protein was reconstituted into phosp hatidylcholine vesicles and its capability to transport phosphate was measu red by phosphate-phosphate exchange. A maximum velocity of 28.1 mu mol min( -1). (mg of protein)(-1) was detected. Phosphate transport was sensitive to mersalyl and N-ethylmaleimide. On the other hand, no proton transport was apparent in these proteoliposomes, unlike to what was seen with Similar hyd roxylapatite-purified fractions obtained after solubilization of the membra ne by decyl-pentaoxyethylene ether, which contained several polypeptides. T hus the phosphate carrier is distinct from the uncoupling protein (UCP) fam ily and its behavior shows that not all anion carriers are able to act as U CPs, (C) 1999 Published by Elsevier Science Ireland Ltd. All rights reserve d.