Mitochondrial Phosphate Carrier: Relation of its SH Groups to Oligomeric Organization

Abstract

Free sulfhydryl groups are a prerequisite for the function of the phosphate transport protein (PTP) of the inner mitochondrial membrane as the translocation process can be inhibited by a large variety of SH-reagents. Sequential inhibition of phosphate transport by reversibly and irreversibly acting thiol reagents revealed a state of the PTP where the transport function was intact yet the protein was protected against irreversible inactivation. On the basis of these experiments it was suggested that the functional unit of PTP carried two sulfhydryl groups accessible from the external side and the two SH groups would by equivalent in the transport function but react with SH reagents sequentially (Fonyó, 1974). However, investigation of the structure of the purified PTP demonstrated that in the 35 kDa polypeptide chain only one single cysteine (out of a total of 6 or 8) reacted with radiolabelled N-ethylmaleimide (NEM) (Kolbe & Wohlrab, 1985). Data obtained in transport studies and by protein structure analysis could be reconciled by suggesting an oligomeric organization of the functional unit of PTP, consisting of two 35 kDa monomers with one essential SH group belonging to each subunit.