Synthetic cation transport peptides: calcium transport across phospholipid membranes

Molecular aspects of peptide-mediated calcium transport are examined through the study of the cation transport properties of a series of synthetic cyclic octapeptides. These peptides, of general structure cyclo[Glu(OR1)-Sar-Gly-(N-R2)Gly]2 (R1 = H or benzyl ester; R2 = cyclohexyl, n-hexyl, or n-decyl) (and an Asp analogue), contain central binding cavities of geometry and dimensions similar to calcium-binding sites in proteins. Transport in Pressman cells ("thick liquid membranes") demonstrated the ionophorous activity of the synthetic peptides; among physiologically abundant cations, the order of selectivity was Ca2+ greater than Na+, K+ much greater than Mg2+. Cation competition studies further showed that cyclo[Glu(OBz)-Sar-Gly-(N-cyclohexyl)Gly]2 (CYCLEX-2E) is essentially a calcium-specific transport peptide whenever calcium is present. When the CYCLEX-2E peptide was added to a suspension of 45Ca2+-loaded sonicated phosphatidylcholine (PC) vesicles in a dialysis sac, the vesicles were completely emptied of internal calcium. Controls using [14C]sucrose established that CYCLEX-2E caused no nonspecific membrane damage. Calcium efflux experiments using several salts of calcium (including 36C1-, [14C]acetate, [14C]succinate, and 35SO4(2-)) suggested that these anions do not specifically accompany the Ca2+-peptide active transporting species across the phospholipid membrane. However, when 45Ca2+-loaded PC vesicles were suspended in mental-free buffer and treated with CYCLEX-2E peptide, calcium efflux did not occur until calcium or sodium chloride was added to the external medium.