A guide to: generation and design of nanobodies

Muyldermans, Serge

doi:10.1111/febs.15515

Cited by 220 publications

(234 citation statements)

References 153 publications

(192 reference statements)

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“…Antibodies associate with high affinity to their cognate target and are very specific to their antigen [ 20 , 55 ]. As antibodies can be raised against virtually any possible molecule, antibodies are already being exploited in the discovery of therapeutics targeting cell plasma membrane transport proteins [ 12 ].…”

Section: Antibodies and Nanobodies To Target Cell Plasma Membrane mentioning

confidence: 99%

“…The HCAbs from camelids recognize antigens by only one single variable domain, known as the variable domain of a H chain of HCAbs (VHH). The VHH fragment, also referred to as nanobody, can be produced recombinantly by a variety of host cells, including, bacteria, yeasts, plants and mammalian cells [ 18 , 19 , 20 ].…”

Section: Antibodies and Nanobodies To Target Cell Plasma Membrane mentioning

confidence: 99%

“…Although nanobodies are the smallest, functional, intact antigen-binding fragments, they are still able to selectively target epitopes selectively and with high affinity. Whereas conventional antibodies and their Fv fragments have a paratope consisting of six CDRs (i.e., three in a VH and three in a VL domain), nanobodies only have three CDRs [ 18 , 19 , 20 ]. Nanobodies are believed to have larger CDRs, more mutation hotspots and recombination signal sequence mimics to compensate for missing VH-VL combinatorial diversity [ 65 , 66 , 67 ].…”

Section: Antibodies and Nanobodies To Target Cell Plasma Membrane mentioning

confidence: 99%

“…Moreover, the smaller size of the footprint and the generally more convex paratope allow nanobodies to target cryptic epitopes, such as the substrate binding site of membrane transport proteins, which are less accessible for conventional antibodies and their derivatives such as the Fab [ 12 , 19 , 61 ]. Furthermore, the single-exon origin (i.e., approximately 360 nucleotides), the intrinsic low immunogenicity, facile blood vessel extravasation, good tissue penetration, robustness upon exposure to extreme conditions and tolerance towards engineering of nanobodies offer advantages for various in vitro and in vivo applications [ 18 , 19 , 20 ].…”

Section: Antibodies and Nanobodies To Target Cell Plasma Membrane mentioning

confidence: 99%

“…The use of conventional antibodies and antigen-binding fragments could overcome some of these issues [ 12 , 13 , 14 , 15 , 16 ]. However, the identification of cell membrane transport protein-targeting antibodies may be limited due to the large size of antibodies and their generally more concave paratope, making small and hidden cryptic epitopes of these proteins potentially unreachable [ 17 , 18 , 19 , 20 ]. Nanobodies are a plausible therapeutic alternative for targeting cell plasma membrane transport proteins.…”

Section: Introductionmentioning

confidence: 99%

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Therapeutic Nanobodies Targeting Cell Plasma Membrane Transport Proteins: A High-Risk/High-Gain Endeavor

et al. 2021

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Cell plasma membrane proteins are considered as gatekeepers of the cell and play a major role in regulating various processes. Transport proteins constitute a subclass of cell plasma membrane proteins enabling the exchange of molecules and ions between the extracellular environment and the cytosol. A plethora of human pathologies are associated with the altered expression or dysfunction of cell plasma membrane transport proteins, making them interesting therapeutic drug targets. However, the search for therapeutics is challenging, since many drug candidates targeting cell plasma membrane proteins fail in (pre)clinical testing due to inadequate selectivity, specificity, potency or stability. These latter characteristics are met by nanobodies, which potentially renders them eligible therapeutics targeting cell plasma membrane proteins. Therefore, a therapeutic nanobody-based strategy seems a valid approach to target and modulate the activity of cell plasma membrane transport proteins. This review paper focuses on methodologies to generate cell plasma membrane transport protein-targeting nanobodies, and the advantages and pitfalls while generating these small antibody-derivatives, and discusses several therapeutic nanobodies directed towards transmembrane proteins, including channels and pores, adenosine triphosphate-powered pumps and porters.

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Section: Antibodies and Nanobodies To Target Cell Plasma Membrane mentioning

confidence: 99%

Section: Antibodies and Nanobodies To Target Cell Plasma Membrane mentioning

confidence: 99%

Section: Antibodies and Nanobodies To Target Cell Plasma Membrane mentioning

confidence: 99%

Section: Antibodies and Nanobodies To Target Cell Plasma Membrane mentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 3 more Smart Citations

Therapeutic Nanobodies Targeting Cell Plasma Membrane Transport Proteins: A High-Risk/High-Gain Endeavor

et al. 2021

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Nanobody engineering: computational modelling and design for biomedical and therapeutic applications

El Salamouni,

Cater,

Spenkelink

et al. 2024

FEBS Open Bio

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Nanobodies, the smallest functional antibody fragment derived from camelid heavy‐chain‐only antibodies, have emerged as powerful tools for diverse biomedical applications. In this comprehensive review, we discuss the structural characteristics, functional properties, and computational approaches driving the design and optimisation of synthetic nanobodies. We explore their unique antigen‐binding domains, highlighting the critical role of complementarity‐determining regions in target recognition and specificity. This review further underscores the advantages of nanobodies over conventional antibodies from a biosynthesis perspective, including their small size, stability, and solubility, which make them ideal candidates for economical antigen capture in diagnostics, therapeutics, and biosensing. We discuss the recent advancements in computational methods for nanobody modelling, epitope prediction, and affinity maturation, shedding light on their intricate antigen‐binding mechanisms and conformational dynamics. Finally, we examine a direct example of how computational design strategies were implemented for improving a nanobody‐based immunosensor, known as a Quenchbody. Through combining experimental findings and computational insights, this review elucidates the transformative impact of nanobodies in biotechnology and biomedical research, offering a roadmap for future advancements and applications in healthcare and diagnostics.

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Development of nanobodies against the coat protein of maize chlorotic mottle virus

Njeru,

Zwaenepoel,

Haesaert

et al. 2024

FEBS Open Bio

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Maize lethal necrosis (MLN) is a maize disease caused by the maize chlorotic mottle virus (MCMV), a potyvirus which causes yield losses of 30–100%. The present study aimed to isolate nanobodies against the MCMV coat protein (CP) for the diagnosis of MLN. MCMV CP expressed in Escherichia coli was used for llama immunization. VHH (i.e. variable heavy domain of heavy chain) gene fragments were prepared from blood drawn from the immunized llama and used to generate a library in E. coli TG1 cells. MCMV specific nanobodies were selected by three rounds of phage display and panning against MCMV CP. The selected nanobodies were finally expressed in E. coli WK6 cells and purified. Eleven MCMV‐specific nanobodies were identified and shown to detect MCMV in infected maize plants. Thus, our results show that nanobodies isolated from llama immunized with MCMV CP can distinguish infected and healthy maize plants, potentially enabling development of affordable MCMV detection protocols.

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A guide to: generation and design of nanobodies

Cited by 220 publications

References 153 publications

Therapeutic Nanobodies Targeting Cell Plasma Membrane Transport Proteins: A High-Risk/High-Gain Endeavor

Therapeutic Nanobodies Targeting Cell Plasma Membrane Transport Proteins: A High-Risk/High-Gain Endeavor

Nanobody engineering: computational modelling and design for biomedical and therapeutic applications

Development of nanobodies against the coat protein of maize chlorotic mottle virus

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