Generation of SUMO-1 modified proteins in E. coli: towards understanding the biochemistry/structural biology of the SUMO-1 pathway.
Here, we developed a binary vector system that introduces a synthetic SUMO-1 conjugation pathway into Escherichia coli and demonstrated that large amounts of sumoylated Ran GTPase activating protein 1 C-terminal region (RanGAP1-C2), Ran binding protein 2 internal repeat domain, p53 and promyelocytic leukemia were efficiently produced. The sumoylated recombinant RanGAP1-C2 appeared to retain the in vivo properties, since it was specifically sumoylated at lysine 517 as expected from in vivo studies. Our findings indicate the establishment of a biosynthetic route for producing large amounts of sumoylated recombinant proteins that will open up new avenues for studying the biochemical and structural aspects of the SUMO-1 modification pathway.
Uchimura
Yasuhiro
Y
Department of Regeneration Medicine, Institute of Molecular Embryology and Genetics, Kumamoto University, 2-2-1 Honjo, Kumamoto 860-0811, Japan.
Nakao
Mitsuyoshi
M
Saitoh
Hisato
H
D000818:Animals; D001665:Binding Sites; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics*; D020690:GTPase-Activating Proteins / Q000096:biosynthesis / Q000378:metabolism; D005822:Genetic Vectors; D051379:Mice; D011499:Protein Processing, Post-Translational; D011994:Recombinant Proteins / Q000096:biosynthesis; D025842:SUMO-1 Protein / Q000235:genetics* / Q000378:metabolism*; D044764:Ubiquitin-Activating Enzymes / Q000235:genetics / Q000378:metabolism
D020690:GTPase-Activating Proteins; D011994:Recombinant Proteins; D025842:SUMO-1 Protein; C470301:UBA2 protein, human; C470299:SAE1 protein, human; D044764:Ubiquitin-Activating Enzymes