Biologically active recombinant human Progastrins(6-80) contains a tightlybound calcium ion

Evidence is accumulating that gastrin precursors may act as growth factors for the colonic mucosa in vivo. The aims of this study were to prepare reco mbinant human progastrin(6-80) and to investigate its structure and biologi cal activities in vitro. Human progastrin(6-80) was expressed in Escherichi a coli as a glutathione S-transferase fusion protein. After thrombin cleava ge progastrine(6-80) was purified by reverse phase high pressure liquid chr omatography and characterized by radioimmunoassay, amino acid sequencing, a nd mass spectrometry, Assays for metal ions by atomic emission spectroscopy revealed the presence of a single tightly bound calcium ion. Progastrin(6- 80) at concentrations in the pM to nM range stimulated proliferation of the conditionally transformed mouse colon cell line YAMC. The observations tha t progastrin(6-80) did not bind to either the cholecystokinin (CCK)-A or th e gastrin/CCK-B receptor expressed in COS cells and that antagonists select ive for either receptor did not reverse the proliferative effects of progas trin(6-80) suggested that progastrin(6-80) stimulated proliferation indepen dently of either the CCK-A or the gastrin/CCK-B receptor. We conclude that recombinant human progastrin(6-80) is biologically active and contains a si ngle calcium ion. With the exception of the well known zinc-dependent polym erization of insulin and proinsulin, this is the first report of selective, high affinity binding of metal ions to a prohormone.