Codon optimization regulates IgG3 and IgM expression and glycosylation in N. benthamiana
Plants are being increasingly recognized for the production of complex human proteins, including monoclonal antibodies (mAbs). Various methods have been applied to boost recombinant expression, with DNA codon usage being an important approach. Here, we transiently expressed three complex human mAbs...
| Main Authors: | , , , , , |
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| Format: | Article |
| Language: | English |
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Frontiers Media S.A.
2023-12-01
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| Series: | Frontiers in Bioengineering and Biotechnology |
| Subjects: | |
| Online Access: | https://www.frontiersin.org/articles/10.3389/fbioe.2023.1320586/full |
| _version_ | 1797403170168635392 |
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| author | Lin Sun Somanath Kallolimath Roman Palt Florian Eminger Richard Strasser Herta Steinkellner |
| author_facet | Lin Sun Somanath Kallolimath Roman Palt Florian Eminger Richard Strasser Herta Steinkellner |
| author_sort | Lin Sun |
| collection | DOAJ |
| description | Plants are being increasingly recognized for the production of complex human proteins, including monoclonal antibodies (mAbs). Various methods have been applied to boost recombinant expression, with DNA codon usage being an important approach. Here, we transiently expressed three complex human mAbs in Nicotiana benthamiana, namely one IgG3 and two IgM directed against SARS-CoV-2 as codon optimized(CO) and non-codon optimized (NCO) variants. qRT-PCR exhibited significantly increased mRNA levels of all CO variants compared to the non-codon optimized orthologues, in line with increased protein expression. Purified CO and NCO mAbs did not exhibit obvious biochemical differences, as determined by SDS-PAGE and antigen binding activities. By contrast, enhanced production selectively impacts on glycosite occupancy and N-glycan processing, with increased mannosidic structures. The results point to a careful monitoring of recombinant proteins upon enhancing expression. Especially if it comes to therapeutic application even subtle modifications might alter product efficacy or increase immunogenicity. |
| first_indexed | 2024-03-09T02:35:34Z |
| format | Article |
| id | doaj.art-8662b700bc6c40acb95eb638acfbc028 |
| institution | Directory Open Access Journal |
| issn | 2296-4185 |
| language | English |
| last_indexed | 2024-03-09T02:35:34Z |
| publishDate | 2023-12-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Bioengineering and Biotechnology |
| spelling | doaj.art-8662b700bc6c40acb95eb638acfbc0282023-12-06T10:33:09ZengFrontiers Media S.A.Frontiers in Bioengineering and Biotechnology2296-41852023-12-011110.3389/fbioe.2023.13205861320586Codon optimization regulates IgG3 and IgM expression and glycosylation in N. benthamianaLin SunSomanath KallolimathRoman PaltFlorian EmingerRichard StrasserHerta SteinkellnerPlants are being increasingly recognized for the production of complex human proteins, including monoclonal antibodies (mAbs). Various methods have been applied to boost recombinant expression, with DNA codon usage being an important approach. Here, we transiently expressed three complex human mAbs in Nicotiana benthamiana, namely one IgG3 and two IgM directed against SARS-CoV-2 as codon optimized(CO) and non-codon optimized (NCO) variants. qRT-PCR exhibited significantly increased mRNA levels of all CO variants compared to the non-codon optimized orthologues, in line with increased protein expression. Purified CO and NCO mAbs did not exhibit obvious biochemical differences, as determined by SDS-PAGE and antigen binding activities. By contrast, enhanced production selectively impacts on glycosite occupancy and N-glycan processing, with increased mannosidic structures. The results point to a careful monitoring of recombinant proteins upon enhancing expression. Especially if it comes to therapeutic application even subtle modifications might alter product efficacy or increase immunogenicity.https://www.frontiersin.org/articles/10.3389/fbioe.2023.1320586/fullcodon usageantibodytranscription and translationglycosylationNicotiana benthamiana |
| spellingShingle | Lin Sun Somanath Kallolimath Roman Palt Florian Eminger Richard Strasser Herta Steinkellner Codon optimization regulates IgG3 and IgM expression and glycosylation in N. benthamiana Frontiers in Bioengineering and Biotechnology codon usage antibody transcription and translation glycosylation Nicotiana benthamiana |
| title | Codon optimization regulates IgG3 and IgM expression and glycosylation in N. benthamiana |
| title_full | Codon optimization regulates IgG3 and IgM expression and glycosylation in N. benthamiana |
| title_fullStr | Codon optimization regulates IgG3 and IgM expression and glycosylation in N. benthamiana |
| title_full_unstemmed | Codon optimization regulates IgG3 and IgM expression and glycosylation in N. benthamiana |
| title_short | Codon optimization regulates IgG3 and IgM expression and glycosylation in N. benthamiana |
| title_sort | codon optimization regulates igg3 and igm expression and glycosylation in n benthamiana |
| topic | codon usage antibody transcription and translation glycosylation Nicotiana benthamiana |
| url | https://www.frontiersin.org/articles/10.3389/fbioe.2023.1320586/full |
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