A Novel Apoptosis Correlated Molecule: Expression and Characterization of Protein Latcripin-1 from Lentinula edodes C91–3

International Journal of Molecular Sciences

Ben Liu¹  Mintao Zhong¹  Yongzhi Lun¹  Xiaoli Wang¹  Wenchang Sun¹  Xingyun Li¹  Anhong Ning¹  Jing Cao¹  Wei Zhang¹  Lei Liu¹ 

[1] Department of Microbiology, Dalian Medical University, No. 9 West Section, Lvshun South Road, Lvshunkou District, Dalian 116044, China;

关键词: Lentinula edodes;    apoptosis;    protein Latcripin-1;    Pichia pastoris;    expression;   

DOI  :  10.3390/ijms13056246

来源: mdpi

【 摘 要 】

An apoptosis correlated molecule—protein Latcripin-1 of Lentinula edodes C_91–3—was expressed and characterized in Pichia pastoris GS115. The total RNA was obtained from Lentinula edodes C_91–3. According to the transcriptome, the full-length gene of Latcripin-1 was isolated with 3′-Full Rapid Amplification of cDNA Ends (RACE) and 5′-Full RACE methods. The full-length gene was inserted into the secretory expression vector pPIC9K. The protein Latcripin-1 was expressed in Pichia pastoris GS115 and analyzed by Sodium Dodecylsulfonate Polyacrylate Gel Electrophoresis (SDS-PAGE) and Western blot. The Western blot showed that the protein was expressed successfully. The biological function of protein Latcripin-1 on A549 cells was studied with flow cytometry and the 3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyl-tetrazolium Bromide (MTT) method. The toxic effect of protein Latcripin-1 was detected with the MTT method by co-culturing the characterized protein with chick embryo fibroblasts. The MTT assay results showed that there was a great difference between protein Latcripin-1 groups and the control group (p < 0.05). There was no toxic effect of the characterized protein on chick embryo fibroblasts. The flow cytometry showed that there was a significant difference between the protein groups of interest and the control group according to apoptosis function (p < 0.05). At the same time, cell ultrastructure observed by transmission electron microscopy supported the results of flow cytometry. The work demonstrates that protein Latcripin-1 can induce apoptosis of human lung cancer cells A549 and brings new insights into and advantages to finding anti-tumor proteins.

【 授权许可】

CC BY   
© 2012 by the authors; licensee Molecular Diversity Preservation International, Basel, Switzerland.