Production of sumoylated proteins using a baculovirus expression system.
Spodoptera frugiperda Sf9 cells were found to possess an active endogenous sumoylation system. However, the endogenous sumoylation machinery did not efficiently modify exogenous proteins expressed by infection with recombinant baculoviruses. To overcome this limitation, mammalian sumoylation components were introduced by co-infection with recombinant baculoviruses expressing individual protein components of the sumoylation pathway. Expression of mammalian Ubc9 plus SUMO (either SUMO1 or SUMO3) was necessary and sufficient for active sumoylation of co-infected test proteins. This system provides a simple and convenient means to produce sumoylated mammalian proteins in a eukaryotic environment. Large-scale cultures should provide quantities of sumoylated proteins sufficient for potential purification.
Langereis
Martijn A
MA
Department of Microbial and Molecular Pathogenesis, College of Medicine, Texas A&M Health Science Center, College Station, TX 77843-1114, USA.
Rosas-Acosta
Germán
G
Mulder
Klaas
K
Wilson
Van G
VG
D000818:Animals; D016367:Baculoviridae / Q000235:genetics; D006224:Cricetinae; D015870:Gene Expression; D005822:Genetic Vectors; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D025842:SUMO-1 Protein / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism; D025841:Small Ubiquitin-Related Modifier Proteins / Q000235:genetics* / Q000378:metabolism; D018411:Spodoptera / Q000821:virology*
D011994:Recombinant Proteins; D025842:SUMO-1 Protein; D025841:Small Ubiquitin-Related Modifier Proteins