Efficient cellular uptake of recombinant murine Hoxc8 homeoprotein in COS-7 cells.
In order to analyze the self-delivery activity of Hoxc8, recombinant Hoxc8 protein (rHoxc8) was designed to be expressed and purified in E. coli as a glutathione S-transferase and green fluorescent protein-fused form (GST-GFP-Hoxc8). After purification using glutathione sepharose beads, the 82 kDa fusion protein was separated on the SDS-PAGE gel and confirmed by detecting the fluorescence through luminescent image analyzer. When rHoxc8 was added to culture media for 30 h, most of the COS-7 cells contained the fusion proteins, showing green fluorescence under the fluorescent microscope. When the efficiency of cellular uptake was examined after Hoechst staining, almost 100% of the cells exhibited the GFP signal, revealing that rHoxc8 can traverse the cellular membrane of COS-7 cells efficiently, suggesting that the rHoxc8 could be applied in the development of efficient and useful delivery vectors for therapeutic molecules.
Lee
Mi Hee
MH
Department of Anatomy, Embryology Lab., Brain Korea 21 Project for Medical Science, Yonsei University College of Medicine, CPO Box 8044, Sodaemoongu Shinchondong 134, Seoul 120-752, Republic of Korea.
Park
Hyoung Woo
HW
Kim
Myoung Hee
MH
D000818:Animals; D019556:COS Cells / Q000378:metabolism; D002522:Chlorocebus aethiops; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005453:Fluorescence; D005982:Glutathione Transferase / Q000235:genetics / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D018398:Homeodomain Proteins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D051379:Mice; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism*; D014161:Transduction, Genetic
D018398:Homeodomain Proteins; C489816:Hoxc8 protein, mouse; D011993:Recombinant Fusion Proteins; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D005982:Glutathione Transferase