Preparation and Properties of Covalently Linked Insulin Dimers

Abstract

The synthesis of 6 isomeric insulin dimers, linked through selected amino groups of the monomers by a dicarboxylic acid, is described. Symmetrical dimers were obtained by direct crosslinking of N,N-bis(methylsulfonylethoxycarbonyl)insulins with the bis(p-nitrophenyl) esters of dicarboxylic acids. The synthesis of asymmetrical dimers was achieved by use of methylsulfonylethoxy carbonyl-protected insulin active ester intermediates. N.epsilon.B29, N.epsilon.B29''-Insulin dimers containing oxalyl, suberoyl and dodecanedioyl crosslinks were produced. N.alpha.B1, N.epsilon.B29''-Insulin dimers with suberoyl and dodecanedioyl crosslinks were synthesized; all other dimers were synthesized with suberoyl crosslinks. The positions of crosslinks were determined by sulfitolysis, tryptic digestion, electrophoresis and quantitative end-group determination. The dimers showed potencies between 1-60% that of insulin on a weight basis in stimulating lipogenesis in isolated fat cells. The potencies are considerably lower than the relative binding affinities determined with isolated fat cells.