Expression and purification of functional human glycogen synthase-1 (hGYS1) in insect cells

May Khanna, Tsuyoshi Imasaki, Vimbai M. Chikwana, Samantha Perez-Miller, Gerald O. Hunter, Amber Mosley, Yuichiro Takagi, Thomas Hurley

Research output: Contribution to journalArticle

5 Citations (Scopus)

Abstract

We have successfully expressed and purified active human glycogen synthase-1 (hGYS1). Successful production of the recombinant hGYS1 protein was achieved by co-expression of hGYS1 and rabbit glycogenin (rGYG1) using the MultiBac baculovirus expression system (BEVS). Functional measurements of activity ratios of hGYS1 in the absence and presence of glucose-6-phosphate and treatment with phosphatase indicate that the expressed protein is heavily phosphorylated. We used mass spectrometry to further characterize the sites of phosphorylation, which include most of the known regulatory phosphorylation sites, as well as several sites unique to the insect cell over-expression. Obtaining large quantities of functional hGYS1 will be invaluable for future structural studies as well as detailed studies on the effects on specific sites of phosphorylation.

Original languageEnglish
Pages (from-to)78-83
Number of pages6
JournalProtein Expression and Purification
Volume90
Issue number2
DOIs
StatePublished - 2013

Fingerprint

Glycogen Synthase
Insects
Phosphorylation
Glucose-6-Phosphate
Baculoviridae
Phosphoric Monoester Hydrolases
Human Activities
Mass Spectrometry
Proteins
Rabbits

Keywords

  • Human glycogen synthase-1
  • Mass spectrometry
  • MultiBac baculovirus expression system
  • Phosphorylation

ASJC Scopus subject areas

  • Biotechnology

Cite this

Expression and purification of functional human glycogen synthase-1 (hGYS1) in insect cells. / Khanna, May; Imasaki, Tsuyoshi; Chikwana, Vimbai M.; Perez-Miller, Samantha; Hunter, Gerald O.; Mosley, Amber; Takagi, Yuichiro; Hurley, Thomas.

In: Protein Expression and Purification, Vol. 90, No. 2, 2013, p. 78-83.

Research output: Contribution to journalArticle

Khanna, May ; Imasaki, Tsuyoshi ; Chikwana, Vimbai M. ; Perez-Miller, Samantha ; Hunter, Gerald O. ; Mosley, Amber ; Takagi, Yuichiro ; Hurley, Thomas. / Expression and purification of functional human glycogen synthase-1 (hGYS1) in insect cells. In: Protein Expression and Purification. 2013 ; Vol. 90, No. 2. pp. 78-83.
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