Čeština
EnglishEfficient expression of Human papillomavirus 16 E7 oncoprotein fused to C-terminus of Tobacco mosaic virus (TMV) coat protein using molecular chaperones in E. coli
Folwarczna J., Moravec T., Plchova H., Hoffmeisterova H., Cerovska N.
PROTEIN EXPRESSION AND PURIFICATION 85: 152-157, 2012
Keywords: bacterial expression, Human papillomavirus, E7 oncoprotein, chaperone
Abstract: The Human papillomavirus 16 (HPV 16) E7 oncoprotein is a promising candidate for development of anti-cancer therapeutic vaccine. We have prepared the expression construct carrying mutagenized E7 oncoprotein fused to the C – terminus of Tobacco mosaic virus (TMV) coat protein via 15 amino acids β sheet linker. The fusion protein was expressed in Escherichia coli MC 1061 cells. We have obtained high level expression, but most of the protein remained in insoluble inclusion bodies. To increase the ratio of soluble protein various molecular chaperones (TF, DnaK-DnaJ-GrpE, GroEL-GroES) were used. The immunological reactivity of expressed recombinant protein was evaluated with anti-E7 and anti-TMV antibodies. The distribution of expressed product during ultracentrifugation on sucrose gradient was studied.
DOI:
Fulltext: contact IEB authors
IEB authors: Noemi Čeřovská, Hana Hoffmeisterová, Tomáš Moravec, Helena Plchová
PROTEIN EXPRESSION AND PURIFICATION 85: 152-157, 2012
Keywords: bacterial expression, Human papillomavirus, E7 oncoprotein, chaperone
Abstract: The Human papillomavirus 16 (HPV 16) E7 oncoprotein is a promising candidate for development of anti-cancer therapeutic vaccine. We have prepared the expression construct carrying mutagenized E7 oncoprotein fused to the C – terminus of Tobacco mosaic virus (TMV) coat protein via 15 amino acids β sheet linker. The fusion protein was expressed in Escherichia coli MC 1061 cells. We have obtained high level expression, but most of the protein remained in insoluble inclusion bodies. To increase the ratio of soluble protein various molecular chaperones (TF, DnaK-DnaJ-GrpE, GroEL-GroES) were used. The immunological reactivity of expressed recombinant protein was evaluated with anti-E7 and anti-TMV antibodies. The distribution of expressed product during ultracentrifugation on sucrose gradient was studied.
DOI:
Fulltext: contact IEB authors
IEB authors: Noemi Čeřovská, Hana Hoffmeisterová, Tomáš Moravec, Helena Plchová