title: Biologically active recombinant human progastrin(6-80) contains a tightly bound calcium ion.
abstract: 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 recombinant human progastrin(6-80) and to investigate its structure and biological activities in vitro. Human progastrin(6-80) was expressed in Escherichia coli as a glutathione S-transferase fusion protein. After thrombin cleavage progastrin(6-80) was purified by reverse phase high pressure liquid chromatography and characterized by radioimmunoassay, amino acid sequencing, and 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 that progastrin(6-80) did not bind to either the cholecystokinin (CCK)-A or the gastrin/CCK-B receptor expressed in COS cells and that antagonists selective for either receptor did not reverse the proliferative effects of progastrin(6-80) suggested that progastrin(6-80) stimulated proliferation independently 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 single calcium ion. With the exception of the well known zinc-dependent polymerization of insulin and proinsulin, this is the first report of selective, high affinity binding of metal ions to a prohormone.
authors: University Department of Surgery, Austin Hospital, Heidelberg, Victoria 3084, Australia. g.baldwin@surgeryaustin.unimelb.edu.au
authors: G S
authors: GS
authors: Baldwin
authors: F
authors: F
authors: Hollande
authors: Z
authors: Z
authors: Yang
authors: Y
authors: Y
authors: Karelina
authors: A
authors: A
authors: Paterson
authors: R
authors: R
authors: Strang
authors: D
authors: D
authors: Fourmy
authors: G
authors: G
authors: Neumann
authors: A
authors: A
authors: Shulkes
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D002118:Calcium / Q000378:metabolism*; D002455:Cell Division / Q000187:drug effects; D005755:Gastrins / Q000737:chemistry* / Q000302:isolation & purification / Q000502:physiology; D006801:Humans; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry*; D011498:Protein Precursors / Q000737:chemistry* / Q000302:isolation & purification / Q000502:physiology; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000494:pharmacology; D012844:Sincalide / Q000378:metabolism
chemical_list: D005755:Gastrins; D010446:Peptide Fragments; D011498:Protein Precursors; D011994:Recombinant Proteins; C013706:big gastrin; D012844:Sincalide; D002118:Calcium