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Related Concept Videos

Antibody Structure01:10

Antibody Structure

Overview
Antibodies, also known as immunoglobulins (Ig), are essential players of the adaptive immune system. These antigen-binding proteins are produced by B cells and make up 20 percent of the total blood plasma by weight. In mammals, antibodies fall into five different classes, which each elicits a different biological response upon antigen binding.
The Y-Shaped Structure of Antibodies Consists of Four Polypeptide Chains
Antibodies consist of four polypeptide chains: two identical heavy...
Antibody Structure01:10

Antibody Structure

Overview
Antibodies, also known as immunoglobulins (Ig), are essential players of the adaptive immune system. These antigen-binding proteins are produced by B cells and make up 20 percent of the total blood plasma by weight. In mammals, antibodies fall into five different classes, which each elicits a different biological response upon antigen binding.
The Y-Shaped Structure of Antibodies Consists of Four Polypeptide Chains
Antibodies consist of four polypeptide chains: two identical heavy...
Antibody Structure and Classes01:25

Antibody Structure and Classes

Antibodies, also known as immunoglobulins, are produced by B cells in response to foreign substances, such as bacteria and viruses. These proteins are critical for recognizing and neutralizing these substances, protecting the body from potential harm.
The basic structure of an antibody consists of four protein chains: two identical heavy chains and two identical light chains. These chains are held together by disulfide bonds and other non-covalent interactions, forming a Y-shaped structure.
Hybridoma Technology01:31

Hybridoma Technology

Hybridoma technology is used for the large-scale production of monoclonal antibodies. Monoclonal antibodies bind to only a single antigenic determinant or epitope. Such antibodies are used in research, diagnostics, and disease therapy. The hybridoma technology established in 1975 by Georges Köhler and Cesar Milstein was awarded the Nobel Prize in Medicine in 1984 for revolutionizing research and therapy.
Hybridoma Selection
Commonly used fusion techniques — electroporation, polyethylene glycol...

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Related Experiment Video

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Bacterial Inner-membrane Display for Screening a Library of Antibody Fragments
12:28

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Published on: October 15, 2016

Engineered human antibody constant domains with increased stability.

Rui Gong1, Bang K Vu, Yang Feng

  • 1Protein Interactions Group, CCRNP, CCR, National Institutes of Health, Frederick, Maryland 21702, USA.

The Journal of Biological Chemistry
|March 25, 2009
PubMed
Summary
This summary is machine-generated.

Engineered immunoglobulin CH2 domains offer enhanced stability for therapeutic development. A novel mutant (m01) exhibits remarkable thermal and chemical stability, making it a promising scaffold for new antibody-based therapies.

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Bacterial Inner-membrane Display for Screening a Library of Antibody Fragments
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Genetic Encoding of a Non-Canonical Amino Acid for the Generation of Antibody-Drug Conjugates Through a Fast Bioorthogonal Reaction
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Laboratory Scale Production and Purification of a Therapeutic Antibody
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Laboratory Scale Production and Purification of a Therapeutic Antibody

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Area of Science:

  • Immunology
  • Protein Engineering
  • Biochemistry

Background:

  • The immunoglobulin (Ig) constant CH2 domain is crucial for antibody effector functions.
  • Isolated CH2 domains are potential scaffolds for creating diverse binders with effector functions.
  • Previous studies indicated limited thermal stability in isolated murine CH2 domains.

Purpose of the Study:

  • To investigate the unfolding mechanisms of isolated human CH2 domains.
  • To enhance the stability of human CH2 domains for therapeutic applications.
  • To develop novel, stable CH2 scaffolds for drug development.

Main Methods:

  • Cloning of human gamma1 CH2 and construction of cysteine mutants.
  • Thermal stability assessment using circular dichroism and differential scanning calorimetry.
  • Chemical stability evaluation using urea-induced unfolding.
  • Structural characterization of a stable mutant (m01) via mass spectrometry and NMR spectroscopy.

Main Results:

  • Human gamma1 CH2 demonstrated higher thermal stability (Tm = 54.1°C) compared to murine CH2.
  • A specific mutant (m01) exhibited significantly increased thermal stability (Tm = 73.8°C) and urea stability (6.8 M vs 4.2 M).
  • The m01 mutant was highly soluble, monomeric, and possessed an additional disulfide bond without altering secondary structure.

Conclusions:

  • Human CH2 domains possess inherent stability at physiological temperatures.
  • The engineered m01 mutant offers superior stability, making it a viable scaffold for therapeutic development.
  • Both wild-type and engineered CH2 domains show promise for creating novel therapeutics against human diseases.