epidermal-growth-factor and imidazole

Immobilized metal affinity chromatography (IMAC) has been widely used for the specific separation of biopolymers. However, leakage of metal ions from IMAC adsorbents is of concern in IMAC. In this study, we designed a novel tridenate bis(5-methyltetrazolium)amine (BMTA) to reduce the leakage of metal ions by improving the affinity to immobilized metal ions. The ligand was bonded onto silica via three-step reaction to prepare a high-performance IMAC stationary phase. The chromatographic behaviors of ribonuclease A, cytochrome c, and lysozyme on the Cu(II)-, Ni(II)-, and Zn(II)-chelated stationary phase were investigated with respect to pH effect and elution with an imidazole gradient. The retention times of these three proteins increased by increasing the pH of the mobile phase but decreased by increasing the concentration of the competitive displacer. The retaining strength of the three proteins on the chelated stationary phase were in the order Cu(II) > Ni(II) > Zn(II). The behavior of these three proteins was consistent with the properties of a typical IMAC. The BMTA ligand exhibited a much stronger affinity for Cu(II) and Ni(II) than iminodiacetic acid (IDA), which is often regarded as a standard tridentate IMAC ligand. Quantum mechanical calculations at the B3LYP/6-31G level were used to image the coordination mode of the protein-metal ions-BMTA complex. In addition, a fused histidine-tagged cecropin b-human epidermal growth factor (CB-EGF) from Escherichia coli crude extract was purified by the Ni(II)-chelated stationary phase, and the purity of the CB-EGF was determined to be at least 90 %. These results suggest that the BMTA ligand may have potential applications in the preparation of therapeutics. Graphical Abstract A novel ligand of tridenate bis(5-methyltetrazolium)amine (BMTA) was designed to reduce the leakage of metal ions from the column in immobolized metal affinity chromatography (IMAC).

Topics: Adsorption; Chromatography, Affinity; Copper; Cytochromes c; Epidermal Growth Factor; Humans; Hydrogen-Ion Concentration; Imidazoles; Imino Acids; Muramidase; Nickel; Quantum Theory; Ribonuclease, Pancreatic; Silicon Dioxide; Tetrazolium Salts; Zinc

It has been reported previously that most human meningiomas have receptors for somatostatin. Here we report the results of investigations of the effect of somatostatin and the somatostatin analog octreotide on the growth in vitro of human meningioma cells. Neither somatostatin nor its analog showed a direct growth inhibitory action on cultured human meningioma cells. Rather, there was a slight but significant stimulation of growth in the presence of somatostatin. The somatostatin receptors in meningioma tissue were shown to be functional since somatostatin inhibited forskolin-stimulated formation of cAMP by meningioma membranes. In addition, cAMP inhibited the growth of cultured meningioma cells. We conclude that the stimulation by somatostatin of the growth of human meningioma cells in vitro is caused by its inhibitory effect on cAMP formation. These results suggest that therapeutic trials of patients with (recurrent) inoperable meningiomas with somatostatin analogs have to be carried out with great caution.

Topics: Adenylyl Cyclase Inhibitors; Bucladesine; Cell Division; Colforsin; Cyclic AMP; DNA Replication; Dose-Response Relationship, Drug; Epidermal Growth Factor; Humans; Imidazoles; Insulin; Kinetics; Meningeal Neoplasms; Meningioma; Octreotide; Receptors, Neurotransmitter; Receptors, Somatostatin; Somatostatin; Thymidine; Tumor Cells, Cultured