TY - JOUR
T1 - Development of Single-Chain Antibodies Specific to Lysophosphatidic Acid Receptor 2
AU - Son, Sang Hyeon
AU - Baek, Seung Il
AU - Ju, Man Seok
AU - Han, Seong Gu
AU - Jung, Sang Taek
AU - Yu, Yeon Gyu
N1 - Funding Information:
We have isolated scFv clones against LPA2using purified P9-LPA2 and an M13 phage library displaying scFv sequence by the biopanning method. The scAbs containing the isolated scFv sequence showed specific binding kinetics to LPA2 with KD values in the nanomolar range. These results suggest that antibodies specific to GPCR can be efficiently obtained using this approach. Determination of the specific binding site of these isolated scAbs in LPA2 and the effect of scAbs on the LPA2-dependent signaling process will be further characterized. Acknowledgments. This research was supported by a grant from National Research Foundation of Korea (2016R1A2B4009952). S. H. Son was supported by a grant from NRF-2017M3A9F6029736 from the National Research Foundation of Korea.
Publisher Copyright:
© 2018 Korean Chemical Society, Seoul & Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
PY - 2018/4
Y1 - 2018/4
N2 - Lysophosphatidic acids (LPAs) are ubiquitous serum phospholipids that trigger diverse cellular responses such as cell proliferation, migration, cell survival, and calcium influx. LPA receptors belong to the G-protein coupled receptor (GPCR) family, and seven subtypes of LPA receptors have been identified. Among them, LPA receptor 2 (LPA2) is involved in the proliferation and metastasis of ovarian, cervical, and breast cancers. Hence, LPA2-specific antagonists or antibodies are considered as potential anticancer therapeutics. To develop antibodies against LPA2, a recombinant LPA2 was expressed in Escherichia coli, purified to homogeneity, and immobilized on a solid surface as an active conformation. An M13 phage library displaying single-chain variable fragments (scFvs) of human IgG containing randomized complementarity-determining regions was applied to select LPA2-specific scFv clones. After five rounds of biopanning, a few scFv clones which showed specific binding to LPA2 were identified. Single-chain antibodies (scAbs), which contain the isolated scFvs and Cκ domain, were constructed and expressed in E. coli. The purified scAbs showed specific binding to LPA2 with KD values of 200–600 nM.
AB - Lysophosphatidic acids (LPAs) are ubiquitous serum phospholipids that trigger diverse cellular responses such as cell proliferation, migration, cell survival, and calcium influx. LPA receptors belong to the G-protein coupled receptor (GPCR) family, and seven subtypes of LPA receptors have been identified. Among them, LPA receptor 2 (LPA2) is involved in the proliferation and metastasis of ovarian, cervical, and breast cancers. Hence, LPA2-specific antagonists or antibodies are considered as potential anticancer therapeutics. To develop antibodies against LPA2, a recombinant LPA2 was expressed in Escherichia coli, purified to homogeneity, and immobilized on a solid surface as an active conformation. An M13 phage library displaying single-chain variable fragments (scFvs) of human IgG containing randomized complementarity-determining regions was applied to select LPA2-specific scFv clones. After five rounds of biopanning, a few scFv clones which showed specific binding to LPA2 were identified. Single-chain antibodies (scAbs), which contain the isolated scFvs and Cκ domain, were constructed and expressed in E. coli. The purified scAbs showed specific binding to LPA2 with KD values of 200–600 nM.
KW - Lysophosphatidic acid receptor 2
KW - P9
KW - Single-chain antibody
KW - Single-chain variable fragment
KW - amphiphilic poly(gamma-glutamate)–ODG
UR - http://www.scopus.com/inward/record.url?scp=85044774012&partnerID=8YFLogxK
U2 - 10.1002/bkcs.11414
DO - 10.1002/bkcs.11414
M3 - Article
AN - SCOPUS:85044774012
SN - 0253-2964
VL - 39
SP - 489
EP - 494
JO - Bulletin of the Korean Chemical Society
JF - Bulletin of the Korean Chemical Society
IS - 4
ER -
