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Adenovirus vector-mediated FAM176A overexpression induces cell death in human H1299 non-small cell lung cancer cells

Xie, Hong;Hu, Jia;Pan, Huan;Lou, Yaxin;Lv, Ping;Chen, Yingyu
- BMB Reports
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- v.47 no.2
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- pp.104-109
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- 2014
FAM176A (family with sequence similarity 176 member A) is a novel molecule related to programmed cell death. A decreased expression of FAM176A has been found in several types of human tumors in including lung cancers. In the present study, we investigated the biological activities of FAM176A on the human non-small cell lung cancer cell line H1299 cells. We constructed a recombinant adenovirus 5-FAM176A vector (Ad5-FAM176A) and evaluated the expression and anti-tumor activities in vitro. Cell viability analysis revealed that the adenovirus-mediated increase of FAM176A inhibited the growth of the tumor cells in a dose- and time-dependent manner. This inhibitory effect was mediated by both autophagy and apoptosis that involved caspase activation. In addition, cell cycle analysis suggested that Ad5-FAM176A could induce cell cycle arrest at the G2/M phase, all of which suggested that adenovirus-mediated FAM176A gene transfer might present a new therapeutic approach for lung cancer treatment.
https://doi.org/10.5483/BMBRep.2014.47.2.090 인용 PDF KSCI KPUBS HTML

Characterization of AprE176, a Fibrinolytic Enzyme from Bacillus subtilis HK176

Jeong, Seon-Ju;Heo, Kyeong;Park, Ji Yeong;Lee, Kang Wook;Park, Jae-Yong;Joo, Sang Hoon;Kim, Jeong Hwan
- Journal of Microbiology and Biotechnology
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- v.25 no.1
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- pp.89-97
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- 2015
Bacillus subtilis HK176 with high fibrinolytic activity was isolated from cheonggukjang, a Korean fermented soyfood. A gene, aprE176, encoding the major fibrinolytic enzyme was cloned from B. subtilis HK176 and overexpressed in E. coli BL21(DE3) using plasmid pET26b(+). The specific activity of purified AprE176 was 216.8 ± 5.4 plasmin unit/mg protein and the optimum pH and temperature were pH 8.0 and 40℃, respectively. Error-prone PCR was performed for aprE176, and the PCR products were introduced into E. coli BL21(DE3) after ligation with pET26b(+). Mutants showing enhanced fibrinolytic activities were screened first using skim-milk plates and then fibrin plates. Among the mutants, M179 showed the highest activity on a fibrin plate and it had one amino acid substitution (A176T). The specific activity of M179 was 2.2-fold higher than that of the wild-type enzyme, but the catalytic efficiency (k_cat/K_m) of M179 was not different from the wild-type enzyme owing to reduced substrate affinity. Interestingly, M179 showed increased thermostability. M179 retained 36% of activity after 5 h at 45℃, whereas AprE176 retained only 11%. Molecular modeling analysis suggested that the 176^th residue of M179, threonine, was located near the cation-binding site compared with the wild type. This probably caused tight binding of M179 with Ca²⁺, whichincreased the thermostability of M179.
https://doi.org/10.4014/jmb.1409.09087 인용 PDF KSCI KPUBS HTML