Highly conserved charge-pair networks in the mitochondrial carrier family

Selection for regain-of-function mutations in the yeast ADP/ATP carrier AAC2 has revealed an unexpected series of charge-pairs. Four of the six amino acids involved are found in the mitochondrial energy transfer motifs used to define this family of proteins. As such, the results found with the ADP/ATP carrier may apply to the family as a whole. Mitochondrial carriers are built from three homologous domains, each with the conserved motif PX(D,E)XX(K,R). Neutralization of the conserved positive charges at K48, R152 or R252 in these motifs results in respiration defective yeast. Neutralization of the negative charges at D149 and D249 also make respiration defective yeast, though E45G or E45Q mutants are able to grow on glycerol. Regain of function occurs when a complementary charge is lost from another site in the molecule. This phenomenon has been observed independently eight times and thus is strong evidence for charge-pairs existing between the affected residues. Five different charge-pairs have been detected in the yeast AAC2 by this method and three more can be predicted based on homology between the domains. The highly conserved charge-pairs occurring within or between the three mitochondrial energy transfer signatures seem to be a critical feature of mitochondrial carrier structure, independent of the substrates transported. Conformational switching between alternative charge-pairs may constitute part of the basis for transport.