TOPOLOGY OF A FUNCTIONALLY IMPORTANT REGION OF THE CARDIAC NA+ CA2+ EXCHANGER/

The cardiac Na+/Ca2+ exchanger, NCX1, has been modeled to consist of 1 1 transmembrane segments and a large cytoplasmic loop (loop f), Cystei ne mutagenesis and sulfhydryl modification experiments demonstrate tha t the loop connecting transmembrane segments 1 and 2 (loop b) is locat ed on the cytoplasmic side of the membrane, as previously modeled. A m utation in loop b, asparagine 101 to cysteine (N101C), renders the exc hanger insensitive to regulation by cytoplasmic Na+ and Ca2+. Nearby m utations at residue threonine 103 (T103C or T103V) increase the appare nt affinity of the exchanger for cytoplasmic Na+ and also produce a si gnificant Li+ transport capacity, The evidence suggests that the regio n at the interface of cytoplasmic loop b and transmembrane segment 2 i s important in Na+ transport and also in secondary regulation, Thus, t his region may form part of the link between the ion translocation pat hway formed by the transmembrane segments and regulatory sites that ha ve previously been localized to loop f.