Abstract
Recombinant butyrylcholinesterase produced in a metabolically regulated transgenic rice cell culture (rrBChE) was purified to produce a highly pure (95%), active form of enzyme. The developed downstream process uses common manufacturing friendly operations including tangential flow filtration, anion-exchange chromatography, and affinity chromatography to obtain a process recovery of 42% active rrBChE. The purified rrBChE was then characterized to confirm its comparability to the native human form of the molecule (hBChE). The recombinant and native enzyme demonstrated comparable enzymatic behavior and had an identical amino acid sequence. However, rrBChE differs in that it contains plant-type complex N-glycans, including an α-1,3 linked core fucose, and a β-1,2 xylose, and lacking a terminal sialic acid. Despite this difference, rrBChE is demonstrated to be an effective stoichiometric bioscavenger for five different organophosphorous nerve agents in vitro. Together, the efficient downstream processing scheme and functionality of rrBChE confirm its promise as a cost-effective alternative to hBChE for prophylactic and therapeutic use.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 1301-1310 |
| Number of pages | 10 |
| Journal | Biotechnology and Bioengineering |
| Volume | 115 |
| Issue number | 5 |
| DOIs | |
| State | Published - May 1 2018 |
Fingerprint
Keywords
- butyrylcholinesterase
- downstream processing
- Hupresin
- N-glycosylation
ASJC Scopus subject areas
- Biotechnology
- Bioengineering
- Applied Microbiology and Biotechnology
Cite this
Purification, characterization, and N-glycosylation of recombinant butyrylcholinesterase from transgenic rice cell suspension cultures. / Corbin, Jasmine M.; Kailemia, Muchena J.; Cadieux, C. Linn; Alkanaimsh, Salem; Karuppanan, Kalimuthu; Rodriguez, Raymond L.; Lebrilla, Carlito B; Cerasoli, Douglas M.; McDonald, Karen A.; Nandi, Somen.
In: Biotechnology and Bioengineering, Vol. 115, No. 5, 01.05.2018, p. 1301-1310.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Purification, characterization, and N-glycosylation of recombinant butyrylcholinesterase from transgenic rice cell suspension cultures
AU - Corbin, Jasmine M.
AU - Kailemia, Muchena J.
AU - Cadieux, C. Linn
AU - Alkanaimsh, Salem
AU - Karuppanan, Kalimuthu
AU - Rodriguez, Raymond L.
AU - Lebrilla, Carlito B
AU - Cerasoli, Douglas M.
AU - McDonald, Karen A.
AU - Nandi, Somen
PY - 2018/5/1
Y1 - 2018/5/1
N2 - Recombinant butyrylcholinesterase produced in a metabolically regulated transgenic rice cell culture (rrBChE) was purified to produce a highly pure (95%), active form of enzyme. The developed downstream process uses common manufacturing friendly operations including tangential flow filtration, anion-exchange chromatography, and affinity chromatography to obtain a process recovery of 42% active rrBChE. The purified rrBChE was then characterized to confirm its comparability to the native human form of the molecule (hBChE). The recombinant and native enzyme demonstrated comparable enzymatic behavior and had an identical amino acid sequence. However, rrBChE differs in that it contains plant-type complex N-glycans, including an α-1,3 linked core fucose, and a β-1,2 xylose, and lacking a terminal sialic acid. Despite this difference, rrBChE is demonstrated to be an effective stoichiometric bioscavenger for five different organophosphorous nerve agents in vitro. Together, the efficient downstream processing scheme and functionality of rrBChE confirm its promise as a cost-effective alternative to hBChE for prophylactic and therapeutic use.
AB - Recombinant butyrylcholinesterase produced in a metabolically regulated transgenic rice cell culture (rrBChE) was purified to produce a highly pure (95%), active form of enzyme. The developed downstream process uses common manufacturing friendly operations including tangential flow filtration, anion-exchange chromatography, and affinity chromatography to obtain a process recovery of 42% active rrBChE. The purified rrBChE was then characterized to confirm its comparability to the native human form of the molecule (hBChE). The recombinant and native enzyme demonstrated comparable enzymatic behavior and had an identical amino acid sequence. However, rrBChE differs in that it contains plant-type complex N-glycans, including an α-1,3 linked core fucose, and a β-1,2 xylose, and lacking a terminal sialic acid. Despite this difference, rrBChE is demonstrated to be an effective stoichiometric bioscavenger for five different organophosphorous nerve agents in vitro. Together, the efficient downstream processing scheme and functionality of rrBChE confirm its promise as a cost-effective alternative to hBChE for prophylactic and therapeutic use.
KW - butyrylcholinesterase
KW - downstream processing
KW - Hupresin
KW - N-glycosylation
UR - http://www.scopus.com/inward/record.url?scp=85042550211&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=85042550211&partnerID=8YFLogxK
U2 - 10.1002/bit.26557
DO - 10.1002/bit.26557
M3 - Article
VL - 115
SP - 1301
EP - 1310
JO - Biotechnology and Bioengineering
JF - Biotechnology and Bioengineering
SN - 0006-3592
IS - 5
ER -