Abstract
Plexins are type I membrane proteins that function as receptors for semaphorins. All of the known plexins contain a large globular domain, termed the sema domain, in the N-terminal extracellular region, which interacts with semaphorins during signal transduction. Here, we describe procedures for protein production and purification that we utilized in the crystallographic study of the mouse Plexin A2 (mPlxnA2) extracellular fragment, including the sema domain. A mutant mammalian cell line, HEK293S GnTI−, was used as an expression host for the production of a crystallizable-quality mPlxnA2 fragment, which contains several N-glycosylation sites and disulfide bonds.
| Original language | English |
|---|---|
| Pages (from-to) | 57-72 |
| Number of pages | 16 |
| Journal | Methods in Molecular Biology |
| Volume | 1493 |
| DOIs | |
| Publication status | Published - 2017 |
Fingerprint
Keywords
- Crystallographic analysis
- Glycosylated protein
- High-density cell culture
- Immunoaffinity purification
- Mammalian expression system
- Stable expression
ASJC Scopus subject areas
- Molecular Biology
- Genetics
Cite this
Immunoaffinity purification of the glycosylated extracellular fragment of mouse Plexin A2 produced in a mammalian expression system. / Nogi, Terukazu; Mihara, Emiko; Yasui, Norihisa; Takagi, Junichi.
In: Methods in Molecular Biology, Vol. 1493, 2017, p. 57-72.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Immunoaffinity purification of the glycosylated extracellular fragment of mouse Plexin A2 produced in a mammalian expression system
AU - Nogi, Terukazu
AU - Mihara, Emiko
AU - Yasui, Norihisa
AU - Takagi, Junichi
PY - 2017
Y1 - 2017
N2 - Plexins are type I membrane proteins that function as receptors for semaphorins. All of the known plexins contain a large globular domain, termed the sema domain, in the N-terminal extracellular region, which interacts with semaphorins during signal transduction. Here, we describe procedures for protein production and purification that we utilized in the crystallographic study of the mouse Plexin A2 (mPlxnA2) extracellular fragment, including the sema domain. A mutant mammalian cell line, HEK293S GnTI−, was used as an expression host for the production of a crystallizable-quality mPlxnA2 fragment, which contains several N-glycosylation sites and disulfide bonds.
AB - Plexins are type I membrane proteins that function as receptors for semaphorins. All of the known plexins contain a large globular domain, termed the sema domain, in the N-terminal extracellular region, which interacts with semaphorins during signal transduction. Here, we describe procedures for protein production and purification that we utilized in the crystallographic study of the mouse Plexin A2 (mPlxnA2) extracellular fragment, including the sema domain. A mutant mammalian cell line, HEK293S GnTI−, was used as an expression host for the production of a crystallizable-quality mPlxnA2 fragment, which contains several N-glycosylation sites and disulfide bonds.
KW - Crystallographic analysis
KW - Glycosylated protein
KW - High-density cell culture
KW - Immunoaffinity purification
KW - Mammalian expression system
KW - Stable expression
UR - http://www.scopus.com/inward/record.url?scp=84994381812&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=84994381812&partnerID=8YFLogxK
U2 - 10.1007/978-1-4939-6448-2_4
DO - 10.1007/978-1-4939-6448-2_4
M3 - Article
C2 - 27787842
AN - SCOPUS:84994381812
VL - 1493
SP - 57
EP - 72
JO - Methods in Molecular Biology
JF - Methods in Molecular Biology
SN - 1064-3745
ER -