title: Simulated microgravity affects semicarbazide-sensitive amine oxidase expression in recombinant Escherichia coli by HPLC-ESI-QQQ analysis.
abstract: Simulated microgravity has been reported to affect the gene, protein expression, and its function in the cells. Semicarbazide-sensitive amine oxidase (SSAO; E.C.1.4.3.6.) is widely distributed in vascular cells, smooth muscle cells, and adipocytes. It is noteworthy whether the expression of SSAO is affected under simulated microgravity or not. In this study, an SSAO-transformed Escherichia coli BL21 was constructed firstly. Then, a sensitive, selective, and accurate method based on high-performance liquid chromatography electrospray ionization triple quadrupole (HPLC-ESI-QQQ) was developed to determine the amount of SSAO in the E. coli BL21. The limit of detection and limit of quantification were 5.0 and 10 fmol, respectively. Finally, SSAO expression in the recombinant E. coli BL21 was evaluated with various gravity and temperature conditions by HPLC-ESI-QQQ analysis. It is interesting that the tendency in the alteration of SSAO under simulated microgravity showed temperature difference. At 18 °C, the amount of SSAO in the inclusion bodies and soluble fractions under the simulated microgravity increased by 83% and 116%, respectively, compared with normal gravity. However, the decrease by 38% and 49% in the inclusion bodies and soluble fractions under the simulated microgravity was observed at 37 °C. Results obtained here indicate that the SSAO expression under simulated microgravity is dramatically sensitive to the temperature. On the other hand, a novel bioreactor from this study may also be useful for the recombinant protein expression in the field of gene engineering.
authors.affiliation: School of Life Science, Beijing Institute of Technology, No.5 Zhongguancun South Street, Haidian District, Beijing, People's Republic of China 100081.
authors.forename: Yongqian
authors.initials: Y
authors.lastname: Zhang
authors.forename: Chengjun
authors.initials: C
authors.lastname: Lai
authors.forename: Jinyan
authors.initials: J
authors.lastname: Duan
authors.forename: Ningxin
authors.initials: N
authors.lastname: Guan
authors.forename: Kaleem
authors.initials: K
authors.lastname: Ullah
authors.forename: Yulin
authors.initials: Y
authors.lastname: Deng
mesh_terms: D006631:Amine Oxidase (Copper-Containing) / Q000096:biosynthesis*; D002623:Chemistry Techniques, Analytical / Q000379:methods; D002851:Chromatography, High Pressure Liquid; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics*; D015870:Gene Expression; D015964:Gene Expression Regulation, Bacterial; D011994:Recombinant Proteins / Q000096:biosynthesis; D021241:Spectrometry, Mass, Electrospray Ionization; D013696:Temperature; D014893:Weightlessness
chemical_list: D011994:Recombinant Proteins; D006631:Amine Oxidase (Copper-Containing)