scholarly journals Kappa-on-Heavy (KoH) bodies are a distinct class of fully-human antibody-like therapeutic agents with antigen-binding properties

2019 ◽  
Vol 117 (1) ◽  
pp. 292-299 ◽  
Author(s):  
Lynn E. Macdonald ◽  
Karoline A. Meagher ◽  
Matthew C. Franklin ◽  
Natasha Levenkova ◽  
Johanna Hansen ◽  
...  
Keyword(s):  
Small Molecules ◽  
Binding Pocket ◽  
Human Antibody ◽  
Antigen Binding ◽  
Wild Type ◽  
Antigen Binding Site ◽  
Binding Properties ◽  
Variable Domains ◽  
Constant Domains ◽  
Ig Heavy Chain

We describe a Kappa-on-Heavy (KoH) mouse that produces a class of highly diverse, fully human, antibody-like agents. This mouse was made by replacing the germline variable sequences of both the Ig heavy-chain (IgH) and Ig kappa (IgK) loci with the human IgK germline variable sequences, producing antibody-like molecules with an antigen binding site made up of 2 kappa variable domains. These molecules, named KoH bodies, structurally mimic naturally existing Bence-Jones light-chain dimers in their variable domains and remain wild-type in their antibody constant domains. Unlike artificially diversified, nonimmunoglobulin alternative scaffolds (e.g., DARPins), KoH bodies consist of a configuration of normal Ig scaffolds that undergo natural diversification in B cells. Monoclonal KoH bodies have properties similar to those of conventional antibodies but exhibit an enhanced ability to bind small molecules such as the endogenous cardiotonic steroid marinobufagenin (MBG) and nicotine. A comparison of crystal structures of MBG bound to a KoH Fab versus a conventional Fab showed that the KoH body has a much deeper binding pocket, allowing MBG to be held 4 Å further down into the combining site between the 2 variable domains.

scholarly journals Hiding in plain sight: structure and sequence analysis reveals the importance of the antibody DE loop for antibody-antigen binding

2020 ◽  
Author(s):  
Simon P. Kelow ◽  
Jared Adolf-Bryfogle ◽  
Roland L. Dunbrack
Keyword(s):  
High Throughput ◽  
High Throughput Sequencing ◽  
Data Bank ◽  
Antigen Binding ◽  
Sequence Variability ◽  
Antigen Binding Site ◽  
Sequencing Studies ◽  
Variable Domains ◽  
Complementarity Determining Regions ◽  
Hiv 1

AbstractAntibody variable domains contain “complementarity determining regions” (CDRs), the loops that form the antigen binding site. CDRs1-3 are recognized as the canonical CDRs. However, a fourth loop sits adjacent to CDR1 and CDR2 and joins the D and E strands on the antibody v-type fold. This “DE loop” is usually treated as a framework region, even though mutations in the loop affect the conformation of the CDRs and residues in the DE loop occasionally contact antigen. We analyzed the length, structure, and sequence features of all DE loops in the Protein Data Bank, as well as millions of sequences from HIV-1 infected and naïve patients. We refer to the DE loop as H4 and L4 in the heavy and light chain respectively. Clustering the backbone conformations of the most common length of L4 (6 residues) reveals four conformations: two κ-only clusters, one λ-only cluster, and one mixed κ/λ cluster. The vast majority of H4 loops are length-8 and exist primarily in one conformation; a secondary conformation represents a small fraction of H4-8 structures. H4 sequence variability exceeds that of the antibody framework in naïve human high-throughput sequences, and both L4 and H4 sequence variability from λ and heavy germline sequences exceed that of germline framework regions. Finally, we identified dozens of structures in the PDB with insertions in the DE loop, all related to broadly neutralizing HIV-1 antibodies, as well as antibody sequences from high-throughput sequencing studies of HIV-infected individuals, illuminating a possible role in humoral immunity to HIV-1.

scholarly journals Mutational landscape of antibody variable domains reveals a switch modulating the interdomain conformational dynamics and antigen binding

2017 ◽  
Vol 114 (4) ◽  
pp. E486-E495 ◽  
Author(s):  
Patrick Koenig ◽  
Chingwei V. Lee ◽  
Benjamin T. Walters ◽  
Vasantharajan Janakiraman ◽  
Jeremy Stinson ◽  
...  
Keyword(s):  
Somatic Mutations ◽  
Conformational Dynamics ◽  
Human Antibody ◽  
Antigen Binding ◽  
Mutational Landscape ◽  
Antibody Maturation ◽  
Binding Function ◽  
Variable Domains

Somatic mutations within the antibody variable domains are critical to the immense capacity of the immune repertoire. Here, via a deep mutational scan, we dissect how mutations at all positions of the variable domains of a high-affinity anti-VEGF antibody G6.31 impact its antigen-binding function. The resulting mutational landscape demonstrates that large portions of antibody variable domain positions are open to mutation, and that beneficial mutations can be found throughout the variable domains. We determine the role of one antigen-distal light chain position 83, demonstrating that mutation at this site optimizes both antigen affinity and thermostability by modulating the interdomain conformational dynamics of the antigen-binding fragment. Furthermore, by analyzing a large number of human antibody sequences and structures, we demonstrate that somatic mutations occur frequently at position 83, with corresponding domain conformations observed for G6.31. Therefore, the modulation of interdomain dynamics represents an important mechanism during antibody maturation in vivo.

scholarly journals Structure of the Anti-C60 Fullerene Antibody Fab Fragment: Structural Determinants of Fullerene Binding

Acta Naturae ◽  
2019 ◽  
Vol 11 (1) ◽  
pp. 58-65 ◽  
Author(s):  
E. M. Osipov ◽  
O. D. Hendrickson ◽  
T. V. Tikhonova ◽  
A. V. Zherdev ◽  
O. N. Solopova ◽  
...  
Keyword(s):  
Binding Site ◽  
Hydrophobic Surface ◽  
Binding Pocket ◽  
C60 Fullerene ◽  
Antigen Binding ◽  
Structural Determinants ◽  
Antigen Binding Site ◽  
Fab Fragment ◽  
X Ray Crystallography ◽  
Solvent Accessible Area

The structure of the anti-C60 fullerene antibody Fab fragment (FabC60) was solved by X-ray crystallography. The computer-aided docking of C60 into the antigen-binding pocket of FabC60 showed that binding of C60 to FabC60 is governed by the enthalpy and entropy; namely, by - stacking interactions with aromatic residues of the antigen-binding site and reduction of the solvent-accessible area of the hydrophobic surface of C60. A fragment of the mobile CDR H3 loop located on the surface of FabC60 interferes with C60 binding in the antigen-binding site, thereby resulting in low antibody affinity for C60. The structure of apo-FabC60 has been deposited with pdbid 6H3H.

scholarly journals The structure of a LAIR1-containing human antibody reveals a novel mechanism of antigen recognition

eLife ◽  
2017 ◽  
Vol 6 ◽  
Author(s):  
Fu-Lien Hsieh ◽  
Matthew K Higgins
Keyword(s):  
Immune Cell ◽  
Adaptive Immune System ◽  
Antigen Recognition ◽  
Binding Potential ◽  
Human Antibody ◽  
Antigen Binding ◽  
Antigen Binding Site ◽  
Fab Fragment ◽  
Cell Recruitment ◽  
Critical Components

Antibodies are critical components of the human adaptive immune system, providing versatile scaffolds to display diverse antigen-binding surfaces. Nevertheless, most antibodies have similar architectures, with the variable immunoglobulin domains of the heavy and light chain each providing three hypervariable loops, which are varied to generate diversity. The recent identification of a novel class of antibody in humans from malaria endemic regions of Africa was therefore surprising as one hypervariable loop contains the entire collagen-binding domain of human LAIR1. Here, we present the structure of the Fab fragment of such an antibody. We show that its antigen-binding site has adopted an architecture that positions LAIR1, while itself being occluded. This therefore represents a novel means of antigen recognition, in which the Fab fragment of an antibody acts as an adaptor, linking a human protein insert with antigen-binding potential to the constant antibody regions which mediate immune cell recruitment.

scholarly journals The crystal structure of avian CD1 reveals a smaller, more primordial antigen-binding pocket compared to mammalian CD1

2008 ◽  
Vol 105 (46) ◽  
pp. 17925-17930 ◽  
Author(s):  
Dirk M. Zajonc ◽  
Harald Striegl ◽  
Christopher C. Dascher ◽  
Ian A. Wilson
Keyword(s):  
Crystal Structure ◽  
Binding Pocket ◽  
Antigen Binding ◽  
Single Chain ◽  
Antigen Binding Site ◽  
Ligand Interaction ◽  
Hydrogen Bond Interactions ◽  
Hydrophobic Binding ◽  
Mhc Locus ◽  
Binding Groove

The molecular details of glycolipid presentation by CD1 antigen-presenting molecules are well studied in mammalian systems. However, little is known about how these non-classical MHC class I (MHCI) molecules diverged from the MHC locus to create a more complex, hydrophobic binding groove that binds lipids rather than peptides. To address this fundamental question, we have determined the crystal structure of an avian CD1 (chCD1–2) that shares common ancestry with mammalian CD1 from ≈310 million years ago. The chCD1–2 antigen-binding site consists of a compact, narrow, central hydrophobic groove or pore rather than the more open, 2-pocket architecture observed in mammalian CD1s. Potential antigens then would be restricted in size to single-chain lipids or glycolipids. An endogenous ligand, possibly palmitic acid, serves to illuminate the mode and mechanism of ligand interaction with chCD1–2. The palmitate alkyl chain is inserted into the relatively shallow hydrophobic pore; its carboxyl group emerges at the receptor surface and is stabilized by electrostatic and hydrogen bond interactions with an arginine residue that is conserved in all known CD1 proteins. In addition, other novel features, such as an A′ loop that interrupts and segments the normally long, continuous α1 helix, are unique to chCD1–2 and contribute to the unusually narrow binding groove, thereby limiting its size. Because birds and mammals share a common ancestor, but the rate of evolution is slower in birds than in mammals, the chCD1–2-binding groove probably represents a more primordial CD1-binding groove.

Application of Molecular Docking for the Development of Improved HIV-1 Reverse Transcriptase Inhibitors

2020 ◽  
Vol 16 ◽  
Author(s):  
Arash Soltani ◽  
Seyed Isaac Hashemy ◽  
Farnaz Zahedi Avval ◽  
Houshang Rafatpanah ◽  
Seyed Abdolrahim Rezaee ◽  
...  
Keyword(s):  
Reverse Transcriptase ◽  
Binding Capacity ◽  
Binding Pocket ◽  
Ligand Docking ◽  
Binding Modes ◽  
Wild Type ◽  
Parent Molecule ◽  
Discovery Studio ◽  
Approved Drugs ◽  
Hiv 1

Introoduction: Inhibition of the reverse transcriptase (RT) enzyme of human immunodeficiency virus (HIV) by low molecular weight inhibitors is still an active area of research. Here, protein-ligand interactions and possible binding modes of novel compounds with the HIV-1 RT binding pocket (the wild-type as well as Y181C and K103N mutants) were obtained and discussed. Methods: A molecular fragment-based approach using FDA-approved drugs were followed to design novel chemical derivatives using delavirdine, efavirenz, etravirine and rilpivirine as the scaffolds. The drug-likeliness of the derivatives was evaluated using Swiss-ADME. Then the parent molecule and derivatives were docked into the binding pocket of related crystal structures (PDB ID: 4G1Q, 1IKW, 1KLM and 3MEC). Genetic Optimization for Ligand Docking (GOLD) Suite 5.2.2 software was used for docking and the results analyzed in the Discovery Studio Visualizer 4. A derivative was chosen for further analysis, if it passed drug-likeliness and the docked energy was more favorable than that of its parent molecule. Out of the fifty-seven derivatives, forty-eight failed in druglikeness screening by Swiss-ADME or in docking stage. Results: The final results showed that the selected compounds had higher predicted binding affinities than their parent scaffolds in both wild-type and the mutants. Binding energy improvement was higher for the structures designed based on second-generation NNRTIs (etravirine and rilpivirine) than the first-generation NNRTIs (delavirdine and efavirenz). For example, while the docked energy for rilpivirine was -51 KJ/mol, it was improved for its derivatives RPV01 and RPV15 up to -58.3 and -54.5 KJ/mol, respectively. Conclusion: In this study, we have identified and proposed some novel molecules with improved binding capacity for HIV RT using fragment-based approach.

scholarly journals Structural and Drug Targeting Insights on Mutant p53

Cancers ◽  
2021 ◽  
Vol 13 (13) ◽  
pp. 3344
Author(s):  
Ana Sara Gomes ◽  
Helena Ramos ◽  
Alberto Inga ◽  
Emília Sousa ◽  
Lucília Saraiva
Keyword(s):  
Small Molecules ◽  
Drug Targeting ◽  
Normal Function ◽  
Structure Stability ◽  
Mutant P53 ◽  
Structural Studies ◽  
Wild Type ◽  
Tumor Onset ◽  
Long Time ◽  
Stability Dynamics

p53 is a transcription factor with a pivotal role in cell homeostasis and fate. Its impairment is a major event in tumor onset and development. In fact, about half of human cancers bear TP53 mutations that not only halt the normal function of p53, but also may acquire oncogenic gain of functions that favor tumorigenesis. Although considered undruggable for a long time, evidence has proven the capability of many compounds to restore a wild-type (wt)-like function to mutant p53 (mutp53). However, they have not reached the clinic to date. Structural studies have strongly contributed to the knowledge about p53 structure, stability, dynamics, function, and regulation. Importantly, they have afforded relevant insights into wt and mutp53 pharmacology at molecular levels, fostering the design and development of p53-targeted anticancer therapies. Herein, we provide an integrated view of mutp53 regulation, particularly focusing on mutp53 structural traits and on targeting agents capable of its reactivation, including their biological, biochemical and biophysical features. With this, we expect to pave the way for the development of improved small molecules that may advance precision cancer therapy by targeting p53.

scholarly journals Semi-Synthesis of Small Molecules of Aminocarbazoles: Tumor Growth Inhibition and Potential Impact on p53

Molecules ◽  
2021 ◽  
Vol 26 (6) ◽  
pp. 1637
Author(s):  
Solida Long ◽  
Joana B. Loureiro ◽  
Carla Carvalho ◽  
Luís Gales ◽  
Lucília Saraiva ◽  
...  
Keyword(s):  
Growth Inhibition ◽  
Small Molecules ◽  
Tumor Cells ◽  
Mutant P53 ◽  
Wild Type ◽  
Naturally Occurring ◽  
Growth Inhibitory ◽  
Growth Inhibitory Activity ◽  
Amino Derivatives ◽  
Carbazole Alkaloids

The tumor suppressor p53 is inactivated by mutation in approximately 50% of human cancers. Small molecules that bind and stabilize those mutants may represent effective anticancer drugs. Herein, we report the tumor cell growth inhibitory activity of carbazole alkaloids and amino derivatives, as well as their potential activation of p53. Twelve aminocarbazole alkaloids were semi-synthesized from heptaphylline (1), 7-methoxy heptaphylline (2), and 7-methoxymukonal (3), isolated from Clausena harmandiana, using a reductive amination protocol. Naturally-occurring carbazoles 1–3 and their amino derivatives were evaluated for their potential effect on wild-type and mutant p53 activity using a yeast screening assay and on human tumor cell lines. Naturally-occurring carbazoles 1–3 showed the most potent growth inhibitory effects on wild-type p53-expressing cells, being heptaphylline (1) the most promising in all the investigated cell lines. However, compound 1 also showed growth inhibition against non-tumor cells. Conversely, semi-synthetic aminocarbazole 1d showed an interesting growth inhibitory activity in tumor cells expressing both wild-type and mutant p53, exhibiting low growth inhibition on non-tumor cells. The yeast assay showed a potential reactivation of mutant p53 by heptaphylline derivatives, including compound 1d. The results obtained indicate that carbazole alkaloids may represent a promising starting point to search for new mutp53-reactivating agents with promising applications in cancer therapy.

scholarly journals The mycoplasma surface proteins MIB and MIP promote the dissociation of the antibody-antigen interaction

2021 ◽  
Vol 7 (10) ◽  
pp. eabf2403
Author(s):  
Pierre Nottelet ◽  
Laure Bataille ◽  
Geraldine Gourgues ◽  
Robin Anger ◽  
Carole Lartigue ◽  
...  
Keyword(s):  
Surface Proteins ◽  
Mycoplasma Species ◽  
Antigen Binding ◽  
Antigen Binding Site ◽  
Fab Fragment ◽  
Cryo Electron Microscopy ◽  
Antigen Complex ◽  
Antigen Interaction ◽  
Immunoglobulin Binding

Mycoplasma immunoglobulin binding (MIB) and mycoplasma immunoglobulin protease (MIP) are surface proteins found in the majority of mycoplasma species, acting sequentially to capture antibodies and cleave off their VH domains. Cryo–electron microscopy structures show how MIB and MIP bind to a Fab fragment in a “hug of death” mechanism. As a result, the orientation of the VL and VH domains is twisted out of alignment, disrupting the antigen binding site. We also show that MIB-MIP has the ability to promote the dissociation of the antibody-antigen complex. This system is functional in cells and protects mycoplasmas from antibody-mediated agglutination. These results highlight the key role of the MIB-MIP system in immunity evasion by mycoplasmas through an unprecedented mechanism, and open exciting perspectives to use these proteins as potential tools in the antibody field.

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