In Planta Glycan Engineering and Functional Activities of IgE Antibodies

Laura Montero-Morales; Daniel Maresch; Silvia Crescioli; Alexandra Castilho; Kristina M. Ilieva; Silvia Mele; Sophia N. Karagiannis; Friedrich Altmann; Herta Steinkellner

doi:10.3389/fbioe.2019.00242

In Planta Glycan Engineering and Functional Activities of IgE Antibodies

Laura Montero-Morales, Daniel Maresch, Silvia Crescioli, Alexandra Castilho, Kristina M. Ilieva, Silvia Mele, Sophia N. Karagiannis, Friedrich Altmann, Herta Steinkellner^*

^*Corresponding author for this work

Dermatology

Research output: Contribution to journal › Article › peer-review

19 Citations (Scopus)

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Abstract

Human immunoglobulin E (IgE) is the most extensively glycosylated antibody isotype so glycans attached to the seven N-glycosites (NGS) in its Fab and Fc domains may modulate its functions. However, targeted modification of glycans in multiply glycosylated proteins remains a challenge. Here, we applied an in vivo approach that allows the manipulation of IgE N-glycans, using a trastuzumab equivalent IgE (HER2-IgE) as a model. Taking advantage of plant inherent features, i.e., synthesis of largely homogeneous complex N-glycans and susceptibility to glycan engineering, we generated targeted glycoforms of HER2-IgE largely resembling those found in serum IgE. Plant-derived HER2-IgE exhibited N-glycans terminating with GlcNAc, galactose or sialic acid, lacking, or carrying core fucose and xylose. We were able to not only modulate the five NGSs naturally decorated with complex N-glycans, but to also induce targeted glycosylation at the usually unoccupied NGS6, thus increasing the overall glycosylation content of HER2-IgE. Recombinant human cell-derived HER2-IgE exhibited large N-glycan heterogeneity. All HER2-IgE variants demonstrated glycosylation-independent binding to the target antigen and the high affinity receptor FcεRI, and subsequent similar capacity to trigger mast cell degranulation. In contrast, binding to the low affinity receptor CD23 (FcεRII) was modulated by the glycan profile, with increased binding to IgE variants with glycans terminating with GlcNAc residues. Here we offer an efficient in planta approach to generate defined glycoforms on multiply glycosylated IgE, allowing the precise exploration of glycosylation-dependent activities.

Original language	English
Article number	242
Pages (from-to)	242
Journal	Frontiers in Bioengineering and Biotechnology
Volume	7
Issue number	SEP
Early online date	25 Sept 2019
DOIs	https://doi.org/10.3389/fbioe.2019.00242
Publication status	Published - 25 Sept 2019

Keywords

Antibodies
Glycan engineering
Glycosylation
IgE
Plants

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10.3389/fbioe.2019.00242

In Planta Glycan Engineering_MONTERO-MORALES_Accepted12Sep2019_GOLD VoRFinal published version, 1.26 MBLicence: CC BY

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title = "In Planta Glycan Engineering and Functional Activities of IgE Antibodies",

abstract = "Human immunoglobulin E (IgE) is the most extensively glycosylated antibody isotype so glycans attached to the seven N-glycosites (NGS) in its Fab and Fc domains may modulate its functions. However, targeted modification of glycans in multiply glycosylated proteins remains a challenge. Here, we applied an in vivo approach that allows the manipulation of IgE N-glycans, using a trastuzumab equivalent IgE (HER2-IgE) as a model. Taking advantage of plant inherent features, i.e., synthesis of largely homogeneous complex N-glycans and susceptibility to glycan engineering, we generated targeted glycoforms of HER2-IgE largely resembling those found in serum IgE. Plant-derived HER2-IgE exhibited N-glycans terminating with GlcNAc, galactose or sialic acid, lacking, or carrying core fucose and xylose. We were able to not only modulate the five NGSs naturally decorated with complex N-glycans, but to also induce targeted glycosylation at the usually unoccupied NGS6, thus increasing the overall glycosylation content of HER2-IgE. Recombinant human cell-derived HER2-IgE exhibited large N-glycan heterogeneity. All HER2-IgE variants demonstrated glycosylation-independent binding to the target antigen and the high affinity receptor FcεRI, and subsequent similar capacity to trigger mast cell degranulation. In contrast, binding to the low affinity receptor CD23 (FcεRII) was modulated by the glycan profile, with increased binding to IgE variants with glycans terminating with GlcNAc residues. Here we offer an efficient in planta approach to generate defined glycoforms on multiply glycosylated IgE, allowing the precise exploration of glycosylation-dependent activities.",

keywords = "Antibodies, Glycan engineering, Glycosylation, IgE, Plants",

author = "Laura Montero-Morales and Daniel Maresch and Silvia Crescioli and Alexandra Castilho and Ilieva, {Kristina M.} and Silvia Mele and Karagiannis, {Sophia N.} and Friedrich Altmann and Herta Steinkellner",

note = "Copyright {\textcopyright} 2019 Montero-Morales, Maresch, Crescioli, Castilho, Ilieva, Mele, Karagiannis, Altmann and Steinkellner.",

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doi = "10.3389/fbioe.2019.00242",

language = "English",

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T1 - In Planta Glycan Engineering and Functional Activities of IgE Antibodies

AU - Montero-Morales, Laura

AU - Maresch, Daniel

AU - Crescioli, Silvia

AU - Castilho, Alexandra

AU - Ilieva, Kristina M.

AU - Mele, Silvia

AU - Karagiannis, Sophia N.

AU - Altmann, Friedrich

AU - Steinkellner, Herta

PY - 2019/9/25

Y1 - 2019/9/25

N2 - Human immunoglobulin E (IgE) is the most extensively glycosylated antibody isotype so glycans attached to the seven N-glycosites (NGS) in its Fab and Fc domains may modulate its functions. However, targeted modification of glycans in multiply glycosylated proteins remains a challenge. Here, we applied an in vivo approach that allows the manipulation of IgE N-glycans, using a trastuzumab equivalent IgE (HER2-IgE) as a model. Taking advantage of plant inherent features, i.e., synthesis of largely homogeneous complex N-glycans and susceptibility to glycan engineering, we generated targeted glycoforms of HER2-IgE largely resembling those found in serum IgE. Plant-derived HER2-IgE exhibited N-glycans terminating with GlcNAc, galactose or sialic acid, lacking, or carrying core fucose and xylose. We were able to not only modulate the five NGSs naturally decorated with complex N-glycans, but to also induce targeted glycosylation at the usually unoccupied NGS6, thus increasing the overall glycosylation content of HER2-IgE. Recombinant human cell-derived HER2-IgE exhibited large N-glycan heterogeneity. All HER2-IgE variants demonstrated glycosylation-independent binding to the target antigen and the high affinity receptor FcεRI, and subsequent similar capacity to trigger mast cell degranulation. In contrast, binding to the low affinity receptor CD23 (FcεRII) was modulated by the glycan profile, with increased binding to IgE variants with glycans terminating with GlcNAc residues. Here we offer an efficient in planta approach to generate defined glycoforms on multiply glycosylated IgE, allowing the precise exploration of glycosylation-dependent activities.

AB - Human immunoglobulin E (IgE) is the most extensively glycosylated antibody isotype so glycans attached to the seven N-glycosites (NGS) in its Fab and Fc domains may modulate its functions. However, targeted modification of glycans in multiply glycosylated proteins remains a challenge. Here, we applied an in vivo approach that allows the manipulation of IgE N-glycans, using a trastuzumab equivalent IgE (HER2-IgE) as a model. Taking advantage of plant inherent features, i.e., synthesis of largely homogeneous complex N-glycans and susceptibility to glycan engineering, we generated targeted glycoforms of HER2-IgE largely resembling those found in serum IgE. Plant-derived HER2-IgE exhibited N-glycans terminating with GlcNAc, galactose or sialic acid, lacking, or carrying core fucose and xylose. We were able to not only modulate the five NGSs naturally decorated with complex N-glycans, but to also induce targeted glycosylation at the usually unoccupied NGS6, thus increasing the overall glycosylation content of HER2-IgE. Recombinant human cell-derived HER2-IgE exhibited large N-glycan heterogeneity. All HER2-IgE variants demonstrated glycosylation-independent binding to the target antigen and the high affinity receptor FcεRI, and subsequent similar capacity to trigger mast cell degranulation. In contrast, binding to the low affinity receptor CD23 (FcεRII) was modulated by the glycan profile, with increased binding to IgE variants with glycans terminating with GlcNAc residues. Here we offer an efficient in planta approach to generate defined glycoforms on multiply glycosylated IgE, allowing the precise exploration of glycosylation-dependent activities.

KW - Antibodies

KW - Glycan engineering

KW - Glycosylation

KW - IgE

KW - Plants

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U2 - 10.3389/fbioe.2019.00242

DO - 10.3389/fbioe.2019.00242

M3 - Article

C2 - 31632959

AN - SCOPUS:85072936757

SN - 2296-4185

VL - 7

SP - 242

JO - Frontiers in Bioengineering and Biotechnology

JF - Frontiers in Bioengineering and Biotechnology

IS - SEP

M1 - 242

ER -

In Planta Glycan Engineering and Functional Activities of IgE Antibodies

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