sdAb-DB: The Single Domain Antibody Database

26Direct control of protein interaction by chemically induced protein proximity (CIPP) holds 27 great potential for cell-and synthetic biology as well as therapeutic applications. However, 28 toxicity, low cell-permeability and lack of orthogonality currently limit the use of available 29 chemical inducers of proximity (CIP). We present 'Mandi', a novel CIP and demonstrate its 30 applicability in cell culture systems as well as living organisms for protein translocation, 31 protein network shuttling and manipulation of endogenous proteins. 32 3 MAIN TEXT 33

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“…12). Such nanobody assisted targeting of chemically induced protein proximity (natCIPP) can easily be extended to other targets 29 .…”

Section: Main Textmentioning

confidence: 99%

A chemical strategy to control protein networks in vivo

Ziegler

Yserentant

Middel

et al. 2020

Preprint

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“…Given the large number of nanobodies that have been selected and characterised 12 , we attempted to expand the LAMA concept to other targets. Nanobodies for G-associated kinase 13 , and for lamina-associated polypeptide 1 14 , did not allow for cpDHFR insertion into the tried positions ( Supplementary Fig.…”

Section: Main Textmentioning

confidence: 99%

Chemogenetic Control of Nanobodies

Farrants

Tarnawski

Müller

et al. 2019

Preprint

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23We introduce an engineered nanobody whose affinity to green fluorescent protein 24 (GFP) can be switched on and off with small molecules. By controlling the cellular 25 localization of GFP fusion proteins, the engineered nanobody allows to study their role 26 in basic biological processes, an approach that should be applicable to numerous 27 previously described GFP fusions. We also outline how the binding affinities of other 28 nanobodies can be controlled by small molecules. 29 30 Supplementary Fig. 1). The most promising insertion hits were in the 55 complementary-determining region 3 (CDR3), which is often essential for making 56 high affinity contacts between nanobodies and their targets 11 . Of these hits, we 57 analyzed GFP LAMAF98, GFP LAMAG97, and GFP LAMAN95 in greater detail (Fig. 1c-e and 58 Supplementary Fig. 2). All three GFP LAMAs retained a single-digit nanomolar affinity 59 to GFP in the absence of ligands. For all three nanobodies, the affinity towards GFP 60 was dramatically decreased in the presence of NADPH and TMP such that no 61 binding to GFP could be detected for GFP LAMAF98 and GFP LAMAG97 (Fig. 1c-e and 62 Supplementary Fig. 3). For GFP LAMAF98 the presence of NADPH alone also 63 affected the binding affinity to GFP, whereas the affinity of GFP LAMAG97 and 64 GFP LAMAN95 was not affected by NADPH. 65

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“…Sequences and structures of synthetic and natural nanobodies can be found in databases. [11,12] Nanobody sequences for specific organisms can be downloaded from the databases and aligned by using several multiple alignment tools available. [13][14][15][16] The alignments can be visualized by specific softwares [17,18], the consensus sequence and the conserved or hypervariable positions can be determined and a template can be obtained similar to the llama-derived nanobody framework proposed previously .…”

Section: Alignmentmentioning

confidence: 99%

“…[13][14][15][16] The alignments can be visualized by specific softwares [17,18], the consensus sequence and the conserved or hypervariable positions can be determined and a template can be obtained similar to the llama-derived nanobody framework proposed previously . [4] Nanobody sequences for Vicugna pacos and Camelus dromedaries are downloaded from single domain Antibody database (sdAb) [11], and aligned separately by using ClustalW, a multiple sequence alignment tool. [16] Alignments are visualized in Unipro U-gene and hypervariable residues, their evolutionary amino acid frequencies and consensus sequences for CDRs are determined.…”

Section: Alignmentmentioning

confidence: 99%

“…Amino acid letter code [11] The positions that retain highly variable in the alignments are labeled as the hypervariable positions and the positions that yield in high gap scores are not considered in the alignment. The amino acid frequencies given in Table 1 and 2 are obtained from the amino acid frequencies found in the hypervariable positions within all aligned sequences for Vicugna pacos and Camelus dromedaries respectively.…”

Section: Camelus Dromedariesmentioning

confidence: 99%

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ModiBodies: A computational method for modifying nanobodies to improve their antigen binding affinity and specificity

Hacisuleyman

Erman

2019

Preprint

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Nanobodies are special derivatives of antibodies, which consist of only a single chain. Their hydrophilic side prevents them from having the solubility and aggregation problems of conventional antibodies, and they retain the similar size and affinity of the binding area to the antigen. Nanobodies have become of considerable interest for next-generation biotechnological tools for antigen recognition. They can be easily engineered due to their high stability and compact size. They have three complementarity determining regions, CDRs, which are enlarged to provide a similar binding surface to that of regular antibodies. The binding residues are more exposed to the environment. One common strategy to improve protein solubility is to replace hydrophobic residues with hydrophilic ones on the binding surface which contributes to both stability and solubility of nanobodies.[1] Here, we propose an algorithm that uses the 3D structures of protein-nanobody complexes as the initial structures and by successive mutations in the CDR domains to find optimum binding amino acids for hypervariable residues of CDRs to increase the binding affinity and nanobody selectivity. We used the MDM4-VH9 complex, (PDB id 2VYR), fructose-bisphosphate aldolase from Trypanosoma congolense, (PDB id 5O0W), and human lysozyme, (PDB id 4I0C). as benchmark studies and identified similar amino acid patterns in hypervariable residues of CDRs with experimentally optimized ones. According to this method, better binding nanobodies can be generated by using this algorithm in a short time. We suggest that this method can complement existing immune and synthetic library-based methods, without the need of experiments or large libraries.

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sdAb-DB: The Single Domain Antibody Database

Cited by 66 publications

References 34 publications

A chemical strategy to control protein networks in vivo

A chemical strategy to control protein networks in vivo

Chemogenetic Control of Nanobodies

ModiBodies: A computational method for modifying nanobodies to improve their antigen binding affinity and specificity

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