AI Article Synopsis

  • * The resulting A20-A26 fusion protein exhibits dramatically enhanced binding affinity and neutralization potency compared to the individual VHs and various control constructs, demonstrating improvements over 330-fold up to 29,000-fold.
  • * Analytical techniques confirm that the A20-A26 fusion binds to the toxin with a 1:1 ratio, effectively engaging both epitopes simultaneously, underscoring the role of molecular geometry in developing powerful antibody agents for targeting antigens.

Article Abstract

Crystal structures of camelid heavy-chain antibody variable domains (VHs) bound to fragments of the combined repetitive oligopeptides domain of toxin A (TcdA) reveal that the C-terminus of VH A20 was located 30 Å away from the N-terminus of VH A26. Based on this observation, we generated a biparatopic fusion protein with A20 at the N-terminus, followed by a (GS) linker and A26 at the C-terminus. This A20-A26 fusion protein shows an improvement in binding affinity and a dramatic increase in TcdA neutralization potency (>330-fold [ ]; ≥2,700-fold [ ]) when compared to the unfused A20 and A26 VHs. A20-A26 also shows much higher binding affinity and neutralization potency when compared to a series of control antibody constructs that include fusions of two A20 VHs, fusions of two A26 VHs, a biparatopic fusion with A26 at the N-terminus and A20 at the C-terminus (A26-A20), and actoxumab. In particular, A20-A26 displays a 310-fold ( ) to 29,000-fold ( ) higher neutralization potency than A26-A20. Size-exclusion chromatography-multiangle light scattering (SEC-MALS) analyses further reveal that A20-A26 binds to TcdA with 1:1 stoichiometry and simultaneous engagement of both A20 and A26 epitopes as expected based on the biparatopic design inspired by the crystal structures of TcdA bound to A20 and A26. In contrast, the control constructs show varied and heterogeneous binding modes. These results highlight the importance of molecular geometric constraints in generating highly potent antibody-based reagents capable of exploiting the simultaneous binding of more than one paratope to an antigen.

Download full-text PDF

Source
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC9909335PMC
http://dx.doi.org/10.3389/fmicb.2023.1110541DOI Listing

Publication Analysis

Top Keywords

neutralization potency
12
a20 a26
12
crystal structures
8
biparatopic fusion
8
fusion protein
8
binding affinity
8
a26 vhs
8
a20
7
a26
7
structure-guided design
4

Similar Publications

Pickering emulsion with tumor vascular destruction and microenvironment modulation for transarterial embolization therapy.

Biomaterials

December 2024

Center of Interventional Radiology and Vascular Surgery, Nurturing Center of Jiangsu Province for State Laboratory of AI Imaging & Interventional Radiology (Southeast University), Department of Radiology, Zhongda Hospital, Medical School, Southeast University, 87 Dingjiaqiao Road, Nanjing, 210009, China; National Innovation Platform for Integration of Medical Engineering Education (NMEE) (Southeast University), Nanjing, 210009, China; Basic Medicine Research and Innovation Center of Ministry of Education, Zhongda Hospital, Southeast University, Nanjing, 210009, China; State Key Laboratory of Digital Medical Engineering, Southeast University, Nanjing, 210009, China. Electronic address:

In the clinic, Lipiodol chemotherapeutic emulsions remain a main choice for patients diagnosed with hepatocellular carcinoma (HCC) via the mini-invasive transarterial chemoembolization (TACE) therapy. However, the poor stability of conventional Lipiodol chemotherapeutic emulsions would result in the fast drug diffusion and incomplete embolization, inducing systemic toxicity and impairing the efficacy of TACE therapy. Therefore, it is of great importance to construct alternative formulations based on commercial Lipiodol to achieve the improved efficacy and safety of HCC treatment.

View Article and Find Full Text PDF

As colorectal cancer (CRC) remains a leading cause of cancer-related death, identifying therapeutic targets and approaches is essential to improve patient outcomes. The EGFR ligand epiregulin (EREG) is highly expressed in RAS wildtype and mutant CRC with minimal expression in normal tissues, making it an attractive target for antibody-drug conjugate (ADC) development. In this study, we produced and purified an EREG monoclonal antibody (mAb), H231, that had high specificity and affinity for human and mouse EREG.

View Article and Find Full Text PDF

In this work, we developed PolyMap (polyclonal mapping), a high-throughput method for mapping protein-protein interactions. We demonstrated the mapping of thousands of antigen-antibody interactions between diverse antibody libraries isolated from convalescent and vaccinated COVID-19 donors and a set of clinically relevant SARS-CoV-2 spike variants. We identified over 150 antibodies with a variety of distinctive binding patterns toward the antigen variants and found a broader binding profile, including targeting of the Omicron variant, in the antibody repertoires of more recent donors.

View Article and Find Full Text PDF

Emerging AXL Inhibitors in Oncology: Chemical and Biological Advances in Targeted Cancer Therapy.

Anticancer Agents Med Chem

December 2024

Institute of Pharmaceutical Research, GLA University, Mathura, UP, 281406, India.

AXL, a receptor tyrosine kinase, has emerged as a critical player in tumorigenesis, metastasis, and resistance to conventional therapies. Its aberrant activation drives cell proliferation, survival, and angiogenesis, making it an attractive target for cancer treatment. In recent years, significant progress has been made in the development of AXL inhibitors.

View Article and Find Full Text PDF

eCD4-Ig is an entry inhibitor of the human immunodeficiency virus (HIV) that mimics the engagement of both CD4 and CCR5 with the HIV Envelope (Env) protein, a property which imbues it with remarkable potency and breadth. However, env is exceptionally genetically malleable and can evolve to escape a wide variety of entry inhibitors. Here we document the evolution of partial eCD4-Ig resistance in simian-HIV (SHIV)-AD8-infected rhesus macaques (RMs) treated with adeno-associated vectors (AAV) encoding eCD4-Ig.

View Article and Find Full Text PDF

Want AI Summaries of new PubMed Abstracts delivered to your In-box?

Enter search terms and have AI summaries delivered each week - change queries or unsubscribe any time!

A PHP Error was encountered

Severity: Notice

Message: fwrite(): Write of 34 bytes failed with errno=28 No space left on device

Filename: drivers/Session_files_driver.php

Line Number: 272

Backtrace:

A PHP Error was encountered

Severity: Warning

Message: session_write_close(): Failed to write session data using user defined save handler. (session.save_path: /var/lib/php/sessions)

Filename: Unknown

Line Number: 0

Backtrace: