Enhanced protein expression in mammalian cells using engineered SUMO fusions: secreted phospholipase A2.
SUMOylation, the covalent attachment of SUMO (small ubiquitin-like modifier), is a eukaryotic post-translational event that has been demonstrated to play a critical role in several biological processes. When used as an N-terminal tag or fusion partner, SUMO has been shown to enhance functional protein production significantly by improving folding, solubility, and stability. We have engineered several SUMOs and, through their fusion, developed a system for enhancing the expression and secretion of complex proteins. To demonstrate the fidelity of this fusion technology, secreted phospholipase A(2) proteins (sPLA(2)) were produced using HEK-293T and CHO-K1 cells. Five mouse sPLA(2) homologs were expressed and secreted in mammalian cell cultures using SUMO or SUMO-derived, N-terminal fusion partners. Mean and median increases of 43- and 18-fold, respectively, were obtained using novel SUMO mutants that are resistant to digestion by endogenous deSUMOylases.
Peroutka
Raymond J
RJ
LifeSensors, Inc., Malvern, Pennsylvania 19355, USA.
Elshourbagy
Nabil
N
Piech
Tara
T
Butt
Tauseef R
TR
D000818:Animals; D016466:CHO Cells; D002460:Cell Line; D006224:Cricetinae; D003412:Cricetulus; D054509:Group X Phospholipases A2 / Q000235:genetics / Q000378:metabolism*; D006639:Histidine / Q000737:chemistry; D006801:Humans; D007668:Kidney / Q000166:cytology; D051379:Mice; D054467:Phospholipases A2 / Q000235:genetics / Q000378:metabolism*; D015202:Protein Engineering / Q000379:methods*; D017510:Protein Folding; D011499:Protein Processing, Post-Translational; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D025841:Small Ubiquitin-Related Modifier Proteins / Q000235:genetics / Q000378:metabolism*; D012995:Solubility
D011993:Recombinant Fusion Proteins; D025841:Small Ubiquitin-Related Modifier Proteins; D006639:Histidine; D054509:Group X Phospholipases A2; D054467:Phospholipases A2; C516366:Pla2g10 protein, mouse