MITOCHONDRIAL ADENINE-NUCLEOTIDE CARRIER - INVESTIGATION OF PRINCIPAL STRUCTURAL, STERIC, AND CONTACT REQUIREMENTS FOR SUBSTRATE BINDING AND TRANSPORT BY MEANS OF RIBOSE-MODIFIED SUBSTRATE-ANALOGS

A selected series of fourteen ribose-modified adenine nucleotide analogues was prepared and characterized as their α-32P- or U-14C-labeled compounds. The capacity of rat liver mitochondria for adenine nucleotide carrier-linked (specific) binding and carrier-mediated transfer across the inner mitochondrial membrane as well as the amount of noncarrier-linked (unspecific) binding of these analogues was determined at 5 °C by means of an inhibitor (atractyloside) stop method and compared with the natural substrates ADP and ATP. Kinetic data of carrier-specific bound analogues were evaluated from Dixon plots and indicate these analogues as competitive inhibitors for mitochondrial [U-14C]AD(T)P uptake. The findings confirm the distinct substrate specificity of the carrier system. By means of the analogues, an experimental proof of the two-step nature of mitochondrial adenine nucleotide translocation, i.e., carrier-specific binding (recognition) and transport, was obtained. Furthermore, the findings provide a detailed description of the basic steric, contact, and structural elements which are prerequisite for carrier-specific binding (A) and additionally for subsequent transport (B): (A) (1) an anti- or syn-positioned β-N-glycosyl-linked purine base; (2) a S- or N-type sugar pucker; (3) a cis disposition of the C(4')-C(5') bond with respect to the heterocycle; (B) (1) a nonfixed anti-positioned purine base with a N-glycosyl torsion angle of approximately -20°; (2) a S-type sugar pucker; (3) a gauche-gauche orientation of the exocyclic C(5')-O(5') group; (4) a trans-positioned, i.e., C(2') ribo hydroxyl, group, which presumably “triggers” the induction of carrier-mediated transport. © 1979, American Chemical Society. All rights reserved.