Immunochemical Approach for Monitoring of Structural Transition of ApoA-I upon HDL Formation Using Novel Monoclonal Antibodies

Abstract Apolipoprotein A-I (apoA-I) undergoes a large conformational reorganization during remodeling of high-density lipoprotein (HDL) particles. To detect structural transition of apoA-I upon HDL formation, we developed novel monoclonal antibodies (mAbs). Splenocytes from BALB/c mice immunized wi...

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Autores principales:	Hitoshi Kimura, Shiho Mikawa, Chiharu Mizuguchi, Yuki Horie, Izumi Morita, Hiroyuki Oyama, Takashi Ohgita, Kazuchika Nishitsuji, Atsuko Takeuchi, Sissel Lund-Katz, Kenichi Akaji, Norihiro Kobayashi, Hiroyuki Saito
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2017
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/cd93be0f7d85464f8c4243caf236703f
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spelling	oai:doaj.org-article:cd93be0f7d85464f8c4243caf236703f2021-12-02T11:53:06ZImmunochemical Approach for Monitoring of Structural Transition of ApoA-I upon HDL Formation Using Novel Monoclonal Antibodies10.1038/s41598-017-03208-82045-2322https://doaj.org/article/cd93be0f7d85464f8c4243caf236703f2017-06-01T00:00:00Zhttps://doi.org/10.1038/s41598-017-03208-8https://doaj.org/toc/2045-2322Abstract Apolipoprotein A-I (apoA-I) undergoes a large conformational reorganization during remodeling of high-density lipoprotein (HDL) particles. To detect structural transition of apoA-I upon HDL formation, we developed novel monoclonal antibodies (mAbs). Splenocytes from BALB/c mice immunized with a recombinant human apoA-I, with or without conjugation with keyhole limpet hemocyanin, were fused with P3/NS1/1-Ag4-1 myeloma cells. After the HAT-selection and cloning, we established nine hybridoma clones secreting anti-apoA-I mAbs in which four mAbs recognize epitopes on the N-terminal half of apoA-I while the other five mAbs recognize the central region. ELISA and bio-layer interferometry measurements demonstrated that mAbs whose epitopes are within residues 1–43 or 44–65 obviously discriminate discoidal and spherical reconstituted HDL particles despite their great reactivities to lipid-free apoA-I and plasma HDL, suggesting the possibility of these mAbs to detect structural transition of apoA-I on HDL. Importantly, a helix-disrupting mutation of W50R into residues 44–65 restored the immunoreactivity of mAbs whose epitope being within residues 44–65 against reconstituted HDL particles, indicating that these mAbs specifically recognize the epitope region in a random coil state. These results encourage us to develop mAbs targeting epitopes in the N-terminal residues of apoA-I as useful probes for monitoring formation and remodeling of HDL particles.Hitoshi KimuraShiho MikawaChiharu MizuguchiYuki HorieIzumi MoritaHiroyuki OyamaTakashi OhgitaKazuchika NishitsujiAtsuko TakeuchiSissel Lund-KatzKenichi AkajiNorihiro KobayashiHiroyuki SaitoNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 7, Iss 1, Pp 1-11 (2017)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Hitoshi Kimura Shiho Mikawa Chiharu Mizuguchi Yuki Horie Izumi Morita Hiroyuki Oyama Takashi Ohgita Kazuchika Nishitsuji Atsuko Takeuchi Sissel Lund-Katz Kenichi Akaji Norihiro Kobayashi Hiroyuki Saito Immunochemical Approach for Monitoring of Structural Transition of ApoA-I upon HDL Formation Using Novel Monoclonal Antibodies
description	Abstract Apolipoprotein A-I (apoA-I) undergoes a large conformational reorganization during remodeling of high-density lipoprotein (HDL) particles. To detect structural transition of apoA-I upon HDL formation, we developed novel monoclonal antibodies (mAbs). Splenocytes from BALB/c mice immunized with a recombinant human apoA-I, with or without conjugation with keyhole limpet hemocyanin, were fused with P3/NS1/1-Ag4-1 myeloma cells. After the HAT-selection and cloning, we established nine hybridoma clones secreting anti-apoA-I mAbs in which four mAbs recognize epitopes on the N-terminal half of apoA-I while the other five mAbs recognize the central region. ELISA and bio-layer interferometry measurements demonstrated that mAbs whose epitopes are within residues 1–43 or 44–65 obviously discriminate discoidal and spherical reconstituted HDL particles despite their great reactivities to lipid-free apoA-I and plasma HDL, suggesting the possibility of these mAbs to detect structural transition of apoA-I on HDL. Importantly, a helix-disrupting mutation of W50R into residues 44–65 restored the immunoreactivity of mAbs whose epitope being within residues 44–65 against reconstituted HDL particles, indicating that these mAbs specifically recognize the epitope region in a random coil state. These results encourage us to develop mAbs targeting epitopes in the N-terminal residues of apoA-I as useful probes for monitoring formation and remodeling of HDL particles.
format	article
author	Hitoshi Kimura Shiho Mikawa Chiharu Mizuguchi Yuki Horie Izumi Morita Hiroyuki Oyama Takashi Ohgita Kazuchika Nishitsuji Atsuko Takeuchi Sissel Lund-Katz Kenichi Akaji Norihiro Kobayashi Hiroyuki Saito
author_facet	Hitoshi Kimura Shiho Mikawa Chiharu Mizuguchi Yuki Horie Izumi Morita Hiroyuki Oyama Takashi Ohgita Kazuchika Nishitsuji Atsuko Takeuchi Sissel Lund-Katz Kenichi Akaji Norihiro Kobayashi Hiroyuki Saito
author_sort	Hitoshi Kimura
title	Immunochemical Approach for Monitoring of Structural Transition of ApoA-I upon HDL Formation Using Novel Monoclonal Antibodies
title_short	Immunochemical Approach for Monitoring of Structural Transition of ApoA-I upon HDL Formation Using Novel Monoclonal Antibodies
title_full	Immunochemical Approach for Monitoring of Structural Transition of ApoA-I upon HDL Formation Using Novel Monoclonal Antibodies
title_fullStr	Immunochemical Approach for Monitoring of Structural Transition of ApoA-I upon HDL Formation Using Novel Monoclonal Antibodies
title_full_unstemmed	Immunochemical Approach for Monitoring of Structural Transition of ApoA-I upon HDL Formation Using Novel Monoclonal Antibodies
title_sort	immunochemical approach for monitoring of structural transition of apoa-i upon hdl formation using novel monoclonal antibodies
publisher	Nature Portfolio
publishDate	2017
url	https://doaj.org/article/cd93be0f7d85464f8c4243caf236703f
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Immunochemical Approach for Monitoring of Structural Transition of ApoA-I upon HDL Formation Using Novel Monoclonal Antibodies

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