ABSTRACT
With IL-6R as target, a new compound 2460A was identified from fungus using HTS screening model. The taxonomics of the produced strain was confirmed to be Trichoderma hazianum rifai after sequencing analysis of rDNA-ITS (internal transcribed spacer). Results showed that this compound has a binding activity on IL-6R competed with IL-6, thus it is a new ligand of IL-6R originating from microbe. With MTT assay, the anti-tumor activities of 2460A were demonstrated on CM126 and HT-29 cell lines separately, the IC50 are 2.17 x 10(-5) mol x L(-1) and 1.8 x 10(-5) mol x L(-1) respectively. The compound affected lightly the HT-29 cell cycle at S phase. Studies for the anti-tumor activity of 2460A in vivo are in progress in our lab.
Subject(s)
Humans , Antineoplastic Agents , Metabolism , Pharmacology , Binding, Competitive , Bone Marrow Neoplasms , Pathology , Cell Cycle , Cell Line, Tumor , Cell Proliferation , HT29 Cells , High-Throughput Screening Assays , Interleukin-6 , Metabolism , Ligands , Receptors, Interleukin-6 , Metabolism , Trichoderma , ChemistryABSTRACT
Many bioactive peptides from neural and endocrine tissue are amidated at C-terminals,which is essential for their activities.The ?-amide comes from post-translational modification that is catalyzed by ?-AE (?-amidating enzyme) or PAM (pepdilylglycine ?-amidating monooxygenase).The gene encoding ?-AE was amplified with PCR and cloned into the plasmid pET-30a.After the recombinant plasmid pET-A was transformed into E.coli BL21,the ?-AE was expressed and purified by the Ni2+affinity chromatography,which has the ability catalyzing Dns-Tyr-Val-Gly to Dns-Tyr-Val-NH2.It identified that the recombinant protein producing by E.coli BL21 is ?-AE,which will benefit for studies of amidation at the C-terminals of peptides.