TY - JOUR
T1 - Negatively charged liposome as a potent inhibitor of post-translation during in vitro synthesis of green fluorescent protein
AU - Bui, Huong Thi
AU - Umakoshi, Hiroshi
AU - Suga, Keishi
AU - Nishida, Masato
AU - Shimanouchi, Toshinori
AU - Kuboi, Ryoichi
N1 - Funding Information:
The fundamental concept of this study was supported by the Research Group of “Membrane Stress Biotechnology.” It was supported in part by a Grant-in-Aid for Scientific Research (No. 15206089, 16686046, 16760635, 17656268, 19656203, 19656220, and 20360350) from the Ministry of Education, Science, Sports, and Culture of Japan, a grant from the 21st Century COE program “Creation of Integrated EcoChemistry” and the Global COE program “Bio-Environmental Chemistry” of the Japan Society for the Promotion of Science (JSPS). The author is also thankful for the useful discussion and comments from Prof. T. Tsuchido, Dr. Y. Matsumura, and Dr. J. Sakamoto (Graduate School of Chemistry, Materials, and Bioengineering, Kansai University, Suita, Japan). The authors are grateful to the Research Center for Solar Energy Chemistry of Osaka University and the Gas hydrate Analyzing System of Osaka University. H.T. Bui acknowledges financial support from the Ministry of Education and Training, Vietnam (MOET) and the Japanese Society of Promotion of Science (JSPS).
PY - 2009/10/1
Y1 - 2009/10/1
N2 - The effect of negatively charged liposome on in vitro synthesis of a reporter protein, green fluorescent protein (GFP), was investigated using a cell-free translation system. GFP was expressed with and without negatively charged liposome prepared with 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphatidylchorine (POPC) and 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoglycerol (POPG), resulting in the GFP fluorescence being reduced to approximately 60% in the presence of POPC/POPG at more than 30% POPG, depending on its concentration. However, the amount of synthesized GFP products, as analyzed by SDS-PAGE, did not change with and without the POPC/POPG liposome. The results of the ultrafiltration operation indicate that the liposome interacts not only with synthesized polypeptide GFP but also with folded synthesized GFP (mature GFP). Liposome also inhibited refolding of unfolded GFP to its native state. The above results show that the POPC/POPG could inhibit the folding of GFP in the post-translational process of the gene expression product.
AB - The effect of negatively charged liposome on in vitro synthesis of a reporter protein, green fluorescent protein (GFP), was investigated using a cell-free translation system. GFP was expressed with and without negatively charged liposome prepared with 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphatidylchorine (POPC) and 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoglycerol (POPG), resulting in the GFP fluorescence being reduced to approximately 60% in the presence of POPC/POPG at more than 30% POPG, depending on its concentration. However, the amount of synthesized GFP products, as analyzed by SDS-PAGE, did not change with and without the POPC/POPG liposome. The results of the ultrafiltration operation indicate that the liposome interacts not only with synthesized polypeptide GFP but also with folded synthesized GFP (mature GFP). Liposome also inhibited refolding of unfolded GFP to its native state. The above results show that the POPC/POPG could inhibit the folding of GFP in the post-translational process of the gene expression product.
KW - Green fluorescent protein (GFP)
KW - Hydrophobicity
KW - Liposome
KW - Protein folding
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U2 - 10.1016/j.bej.2009.05.002
DO - 10.1016/j.bej.2009.05.002
M3 - Article
AN - SCOPUS:67651111837
SN - 1369-703X
VL - 46
SP - 154
EP - 160
JO - Biochemical Engineering Journal
JF - Biochemical Engineering Journal
IS - 2
ER -