CDR1 composition can affect nanobody recombinant expression yields

Orlando, Marco (author), Fortuna, Sara (author), Oloketuyi, Sandra (author), Bajc, Gregor (author), Goldenzweig, Adi (author), De Marco, Ario (author)

The isolation of nanobodies from pre-immune libraries by means of biopanning is a straightforward process. Nevertheless, the recovered candidates often require optimization to improve some of their biophysical characteristics. In principle, CDRs are not mutated because they are likely to be part of the antibody paratope, but in this work, we describe a mutagenesis strategy that specifically addresses CDR1. Its sequence was identified as an instability hot spot by the PROSS program, and the available structural information indicated that four CDR1 residues bound directly to the antigen. We therefore modified the loop flexibility with the addition of an extra glycine rather than by mutating single amino acids. This approach significantly increased the nanobody yields but traded-off with moderate affinity loss. Accurate modeling coupled with atomistic molecular dynamics simulations enabled the modifications induced by the glycine insertion and the rationale behind the engineering design to be described in detail.

• Language: English
• Keywords: nanobodies, in silico engineering, mutagenesis, nanobody CDRs, in silico modeling, rational mutagenesis, nanobody engineering
• Work type: Article
• Typology: 1.01 - Original Scientific Article
• Organization: BF - Biotechnical Faculty
• Publication status: Published
• Publication version: Version of Record
• Year: 2021
• Number of pages: 13 str.
• Numbering: Vol. 11, iss. 9, art. 1362
• PID: 20.500.12556/RUL-136353
• UDC: 577
• ISSN on article: 2218-273X
• DOI: 10.3390/biom11091362
• COBISS.SI-ID: 76397059
• Publication date in RUL: 25.04.2022

Record is a part of a journal

• Title: Biomolecules
• Shortened title: Biomolecules
• Publisher: MDPI
• ISSN: 2218-273X
• COBISS.SI-ID: 519952921

Licences

• License: CC BY 4.0, Creative Commons Attribution 4.0 International
• Link: http://creativecommons.org/licenses/by/4.0/
• Description: This is the standard Creative Commons license that gives others maximum freedom to do what they want with the work as long as they credit the author.
• Licensing start date: 14.09.2021