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Generation of Expression Clone Set for Functional Proteomics of Human Gastric and Liver Cancers

Authors :
Yong Won Park
Nam-Soon Kim
So-Young Jeong
Jung-Hwa Oh
Yeo-Jin Jeon
Nang-Soo Oh
Ji-Seon Park
Jin-Ok Yang
Hyang-Sook Yoo
Source :
Experimental Biology and Medicine. 234:1220-1229
Publication Year :
2009
Publisher :
SAGE Publications, 2009.

Abstract

Two thousand sixty-eight multi-purpose expression clones for the 326 candidate genes related to gastric or liver cancers were constructed using the Gateway system. These clones can be expressed as His, Glutathione-S-transferase (GST) or Enhanced version of the green fluorescent protein (EGFP) fusion proteins in E. coli, insect cells or mammalian cells. For the 246 E. coli expression clones, the GST fusion proteins had greater expression efficiency and solubility than the His fusion proteins. Approximately 20% of the expressed proteins had unexpected molecular weights. A detailed sequence analysis of these clones revealed frameshift mutations resulting from insertion, deletion or substitution of nucleotides. The results indicate that these changes in the candidate genes may affect the occurrence of gastric or liver cancers. In addition, when 105 proteins, which were expressed in E. coli at very low or undetectable levels, were expressed in insect cells, 76% of the proteins were expressed very well and most were soluble. We also found that most of the 30 proteins prepared using EGFP mammalian expression clones were localized to cellular compartments expected by Gene ontology (GO) and this localization was unaffected if the EGFP-fusion was at the N-terminal or C-terminal region of the protein. Antibody production and subcellular localization analysis of the candidate genes as well as a screen of genes involved in carcinogenesis pathways are currently in progress using these expression clones. These studies provide a valuable resource for developing a better understanding of the molecular mechanism of carcinogenesis in both gastric and liver cancer and would be very helpful in diagnosis and therapeutic predictions.

Details

ISSN :
15353699 and 15353702
Volume :
234
Database :
OpenAIRE
Journal :
Experimental Biology and Medicine
Accession number :
edsair.doi.dedup.....9d328ba647bc8e16d1216e6ebd240d2d
Full Text :
https://doi.org/10.3181/0812-rm-371