{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,10,26]],"date-time":"2025-10-26T20:59:12Z","timestamp":1761512352126},"reference-count":15,"publisher":"Oxford University Press (OUP)","issue":"4","license":[{"start":{"date-parts":[[2007,4,1]],"date-time":"2007-04-01T00:00:00Z","timestamp":1175385600000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/academic.oup.com\/journals\/pages\/open_access\/funder_policies\/chorus\/standard_publication_model"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"published-print":{"date-parts":[[2007,4,1]]},"abstract":"<jats:title>Abstract<\/jats:title>\n               <jats:p>We generated three functionally unique monoclonal antibodies to the purified human CaR extracellular domain. Flow cytometry studies of chimeric receptors localized their epitopes to lobe 2 of the VFT domain. These results lead us to propose a mechanism for the functional effects of these antibodies.<\/jats:p>\n               <jats:p>Introduction: The human Ca2+ receptor (CaR), which plays a central role in the regulation of [Ca2+]0 homeostasis, has a distinctively large extracellular domain that consists of a bilobed Venus flytrap (VFT) domain, involved in agonist binding, and a cysteine-rich domain. Functional antibodies that specifically bind to this domain would have therapeutic potential and could be used as a tool to gain insights into receptor activation as well.<\/jats:p>\n               <jats:p>Materials and Methods: We generated three monoclonal antibodies (mAbs), 7F8, 5C8, and 1A8, to the purified human CaR extracellular domain. Functional characterization of these antibodies included Ca2+ stimulation of phosphoinositide hydrolysis to examine effects of intact or protease digested antibodies on sensitivity of the receptor to extracellular Ca2+ and flow cytometry assay of binding of the antibodies to HEK-293 cells expressing chimeric receptors to map antibody epitopes.<\/jats:p>\n               <jats:p>Results: We found these mAbs specifically recognize native but not denatured human CaR or homologous native Fugu CaR. Sensitivity of the human CaR to extracellular calcium was increased by binding of 5C8 but decreased by binding of 1A8. A chimeric receptor FCFCF, with lobe 2 region of the human CaR VFT domain in the Fugu CaR backbone, bound all three mAbs, and the sensitivity of this chimeric CaR to extracellular Ca2+ was also increased by binding of 5C8 and decreased by binding of 1A8.<\/jats:p>\n               <jats:p>Conclusions: The epitopes of these mAbs reside in the lobe 2 region of the human CaR VFT domain. 5C8 might activate the receptor by facilitating closure and\/or rotation of the VFT domains on agonist binding, whereas 1A8 might inhibit the receptor by impeding such agonist-induced conformational changes. Recombinant antibodies with antigen binding domains of 5C8 and 1A8 could be useful in the treatment of hyperparathyroidism and osteoporosis, respectively.<\/jats:p>","DOI":"10.1359\/jbmr.070111","type":"journal-article","created":{"date-parts":[[2007,1,23]],"date-time":"2007-01-23T13:18:51Z","timestamp":1169558331000},"page":"601-608","source":"Crossref","is-referenced-by-count":10,"title":["Functional Effects of Monoclonal Antibodies to the Purified Amino-Terminal Extracellular Domain of the Human Ca2+ Receptor"],"prefix":"10.1093","volume":"22","author":[{"given":"Jianxin","family":"Hu","sequence":"first","affiliation":[{"name":"Molecular Pathophysiology Section, NIDCD, National Institutes of Health, Bethesda, Maryland, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Guadalupe","family":"Reyes-Cruz","sequence":"additional","affiliation":[{"name":"Molecular Pathophysiology Section, NIDCD, National Institutes of Health, Bethesda, Maryland, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Paul K","family":"Goldsmith","sequence":"additional","affiliation":[{"name":"Molecular Pathophysiology Section, NIDCD, National Institutes of Health, Bethesda, Maryland, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Nicole M","family":"Gantt","sequence":"additional","affiliation":[{"name":"Molecular Medicine Branch, NIDDK, National Institutes of Health, Bethesda, Maryland, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Jeffery L","family":"Miller","sequence":"additional","affiliation":[{"name":"Molecular Medicine Branch, NIDDK, National Institutes of Health, Bethesda, Maryland, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Allen M","family":"Spiegel","sequence":"additional","affiliation":[{"name":"Molecular Pathophysiology Section, NIDCD, National Institutes of Health, Bethesda, Maryland, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"286","published-online":{"date-parts":[[2009,12,4]]},"reference":[{"key":"2024020319313775400_bib1","doi-asserted-by":"crossref","first-page":"575","DOI":"10.1038\/366575a0","article-title":"Cloning and characterization of an extracellular Ca2+-sensing receptor from bovine parathyroid","volume":"366","author":"Brown","year":"1993","journal-title":"Nature"},{"key":"2024020319313775400_bib2","doi-asserted-by":"crossref","first-page":"239","DOI":"10.1152\/physrev.2001.81.1.239","article-title":"Extracellular calcium sensing and extracellular calcium signaling","volume":"81","author":"Brown","year":"2001","journal-title":"Physiol Rev"},{"key":"2024020319313775400_bib3","doi-asserted-by":"crossref","first-page":"1723","DOI":"10.1093\/emboj\/18.7.1723","article-title":"Molecular tinkering of G protein-coupled receptors: An evolutionary success","volume":"18","author":"Bockaert","year":"1999","journal-title":"EMBO J"},{"key":"2024020319313775400_bib4","doi-asserted-by":"crossref","first-page":"971","DOI":"10.1038\/35039564","article-title":"Structural basis of glutamate recognition by a dimeric metabotropic glutamate receptor","volume":"407","author":"Kunishima","year":"2000","journal-title":"Nature"},{"key":"2024020319313775400_bib5","doi-asserted-by":"crossref","first-page":"282","DOI":"10.1016\/S1043-2760(03)00104-8","article-title":"Naturally occurring mutations of the extracellular Ca2+-sensing receptor: Implications for its structure and function","volume":"14","author":"Hu","year":"2003","journal-title":"Trends Endocrinol Metab"},{"key":"2024020319313775400_bib6","doi-asserted-by":"crossref","first-page":"11303","DOI":"10.1074\/jbc.274.16.11303","article-title":"Expression, purification, and biochemical characterization of the amino-terminal extracellular domain of the human calcium receptor","volume":"274","author":"Goldsmith","year":"1999","journal-title":"J Biol Chem"},{"key":"2024020319313775400_bib7","doi-asserted-by":"crossref","first-page":"3","DOI":"10.1016\/0076-6879(81)73054-4","article-title":"Preparation of monoclonal antibodies: Strategies and procedures","volume":"73","author":"Galfre","year":"1981","journal-title":"Methods Enzymol"},{"key":"2024020319313775400_bib8","doi-asserted-by":"crossref","first-page":"31355","DOI":"10.1074\/jbc.272.50.31355","article-title":"The carboxyl terminus of the human calcium receptor. 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