Enhanced accumulation of secreted human growth hormone by transgenic tobacco cells correlates with the introduction of an N-glycosylation site |
| |
Authors: | Xu Jianfeng Kieliszewski Marcia |
| |
Institution: | a Department of Chemistry and Biochemistry, Ohio University, Athens, OH 45701, United States b Arkansas Biosciences Institute, College of Agriculture and Technology, Arkansas State University, Jonesboro, AR 72401, United States |
| |
Abstract: | Extracellular secretion of recombinant proteins from plant cell suspension culture will simplify the protein purification procedure and greatly reduce the production cost. Our early work indicated that presence of hydroxyproline-O-glycosylation at the C- or N-terminus of the target protein boosted the secreted yields in the culture medium. Inspired by early successes, we tested the possibility of introducing an N-glycosylation site to facilitate the secretion of human growth hormone (hGH) from cultured tobacco cells. Three N-glycosylated hGH fusion proteins, designated NAS-EK-hGH, NAS-Kex2-hGH and hGH-NAS, were expressed in tobacco BY-2 cells. Where NAS denotes the “Asn-Ala-Ser” consensus sequence for N-glycosylation; EK denotes an enterokinase cleavage site and Kex2 a sequence to be cleaved by a Golgi-localized Kex2p-like protease. Our results indicated that a single N-glycan attached either at the N-terminus or C-terminus of hGH correlated with enhanced extracellular accumulation of the transgenic proteins; the secreted yield of NAS-EK-hGH and hGH-NAS was 70-90 fold greater than the control targeted, non-glycosylated hGH. NAS-Kex2-hGH was subject to partial cleavage of the N-glycan tag at the Kex2 site in Golgi apparatus, and therefore gave lower yields than the other two constructs. |
| |
Keywords: | Plant cell culture Recombinant protein Human growth hormone Glycosylation Secretion |
本文献已被 ScienceDirect PubMed 等数据库收录! |
|