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Study of a Novel Antiosteoporosis Screening Model Targeted on Cathepsin K

Biomedical and Environmental Sciences 2004年第3期17卷 273-280页

作者： JUNYANG GUANG-DONGSHANG yue-qinzhang InstituteofMedicinalBiotechnology ChineseAcademyofMedicalSciencesandPekingUnionMedicalCollegeBeijing100050China DepartmentofChemistryandChemicalBiology CornellUniversityUSA

Objective To establish an effective assay to access the effects of natural products on cathepsin K for screening antiosteoporosis drugs. Methods To obtain the purified cathepsin K, we cloned the target fragment from the mRNA of human osteosacoma cell line MG63 and demonstrated its correctness through DNA sequencing. Cathepsin K was expressed in a high amount in *** after IPTG induction, and was purified to near homogenetity through resolution and column purification. The specificity of the protein was shown by Western blotting experiment. The biological activity of the components in the fermentation broth was assayed by their inhibitory effects on cathepsin K and its analog papain. Results With the inhibition of papain activity as a screen index, the fermentation samples of one thousand strains of fungi were tested and 9 strains among them showed strong inhibitory effects. The crude products of the fermentation broth were tested for their specific inhibitory effects on the purified human cathepsin K, the product of fungi 2358 shows the highest specificity against cathepsin K. Conclusions The compounds isolated from fungi 2358 show the highest biological activity and are worth further structure elucidation and function characterization.

关键词： Cathepsin K Inhibitor Antiosteoporosis Screening model

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Identification of a New Peptide Deformylase Gene From Enterococcus faecium and Establishment of a New Screening Model Targeted on PDF for Novel Antibiotics

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Biomedical and Environmental Sciences 2004年第3期17卷 350-358页

作者： XIAN-BINGTANG SHU-YISI yue-qinzhang TheNationalKeyLaboratoryforScreeningNewMicrobialDrugs InstituteofMedicinalBiotechnologyChineseAcademyofMedicalSciencesandPekingUnionMedicalCollegeBeifing100050China InstituteofMedicinalBiotechnology ChineseAcademyofMediicalSciences＆PekingUnionMedicalCollegeNo.1TiantanXiliBeijing100050China

Objective To identify a new peptide deformylase (PDF) gene (Genebank Accession AY238515) from Enterococcus faecium and to establish a new screening model targeted on PDF. Methods A new PDF gene was identified by BLAST analysis and PCR and was subsequently over-expressed in the prokaryotic expression host *** Bl21(DE3). Over-expressed protein was purified for enzymatic assay by metal affinity chromatography and a new screening model was established for novel antibiotics. Result A new PDF gene of Enterococcus faecium was identified successfully. Ten positive samples were picked up from 8000 compound library and the microbial fermentation broth samples. Conclusion A new PDF of gene Enterococcus faecium was first identified and the model had a high efficacy. Positive samples screened may be antibacterial agents of broad spectrum.

关键词： Enterococcus faecium Peptide deformylase (PDF) Antibacterial gents of broad spectrum

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