PMID: 8939465Nov 1, 1996Paper

Expression, purification and characterization of recombinant C. Elegans UNC-18

Neurochemistry International
H OgawaR Hosono

Abstract

The Caenorhabditis elegans unc-18-encoded protein (UNC-18) is implicated in the processes of vesicle targeting, docking, and/or fusion. To further characterize the properties of this important neural protein, we expressed it at a high level in Spodoptera frugiperda Sf21 cells using a baculovirus expressing system. A cDNA containing the coding sequence for UNC-18 was inserted into the transfer vector pBlueBac to yield the recombinant virus pAcNPV/unc-18. At maximal expression, the recombinant virus produces a protein of 67 kDa, which constitutes about one-third of total cell protein. The UNC-18 protein was highly purified and its biochemical and functional properties were assessed. The protein is globular with an isoelectric point of 6.95. Circular dichroism spectroscopy indicated that the alpha-helix and beta-sheet account for 10.0 and 59.0%, respectively. Immunolabeling the Sf21 cells expressing UNC-18 showed that the expressed UNC-18 is predominantly localized in the cytoplasm as a soluble monomer. The protein is phosphorylated by protein kinase C and binds to the recombinant C. elegans syntaxin in vitro. These findings suggest that in vesicle traffic UNC-18 is a regulator factor associated with the plasma membrane through sy...Continue Reading

References

Jan 11, 1984·Nucleic Acids Research·J DevereuxO Smithies
May 5, 1982·Journal of Molecular Biology·J Kyte, R F Doolittle
Jan 6, 1981·Biochemistry·S W Provencher, J Glöckner
Mar 9, 1994·Annals of the New York Academy of Sciences·G SchiavoC Montecucco
Feb 15, 1994·Proceedings of the National Academy of Sciences of the United States of America·J PevsnerR H Scheller
Mar 15, 1994·Proceedings of the National Academy of Sciences of the United States of America·E P GarciaP De Camilli
Nov 1, 1996·Neurochemistry International·T SassaR Hosono

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Citations

Nov 1, 1996·Neurochemistry International·T SassaR Hosono
Oct 11, 2003·Molecular and Biochemical Parasitology·Sankale Shompole, Douglas P Jasmer
Nov 5, 1999·The Journal of Biological Chemistry·K YamadaT Okazaki
Jan 27, 1998·The Journal of Biological Chemistry·H OgawaR Hosono

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