Diphtheria toxin (DT) is a highly potent protein secreted by the bacterium Corynebacterium diphtheriae, the pathogen responsible for diphtheria. The toxin’s production is governed by the tox gene carried by a specific bacterial virus, known as a bacteriophage (e.g., phage Beta). This genetic transfer allows only lysogenized, or infected, bacterial strains to produce the toxin that causes systemic symptoms. Diphtheria toxin’s effect on human physiology stems from its precise, multi-step molecular mechanism designed to penetrate host cells and disable their core machinery.
The Architecture of Diphtheria Toxin
Diphtheria toxin is initially synthesized as a single polypeptide chain, functioning as a classic A-B type toxin with two primary functional components. The full protein chain is proteolytically processed, or “nicked,” into two main fragments linked by a disulfide bond. This cleavage yields Fragment A (the amino-terminal portion) and Fragment B (the carboxyl-terminal section).
Fragment A contains the Catalytic (C) domain, which holds the enzymatic activity responsible for the toxin’s effects inside the cell. Fragment B is structurally more complex, containing two distinct domains that facilitate the toxin’s entry into the host cell: the Translocation (T) domain and the Receptor-binding (R) domain. The arrangement of these three domains (C, T, and R) ensures the catalytic payload is delivered to the correct intracellular location.
Host Cell Recognition and Internalization
The initial step is the toxin’s binding to the surface of a susceptible host cell, mediated by the Receptor-binding (R) domain of Fragment B. The cellular receptor is the Heparin-binding epidermal growth factor precursor (HB-EGF), a protein found on various cell membranes. This specificity causes the toxin to primarily target tissues like the heart and nerves, which express high levels of this receptor.
Once the R domain binds to HB-EGF, the receptor-toxin complex is engulfed by the host cell through receptor-mediated endocytosis. The complex is enclosed within an endosome, which begins to acidify as proton pumps lower the internal pH, often to a range of 5.0 to 5.5. This drop in pH triggers a structural change in the toxin.
The acidic environment causes the Translocation (T) domain within Fragment B to undergo a conformational change, exposing hydrophobic regions. This shift allows the T domain to insert into the endosomal membrane, forming a channel or pore. The low pH also facilitates the dissociation of the toxin from the HB-EGF receptor. This channel provides the pathway for the catalytic Fragment A to escape the endosome and enter the host cell’s cytoplasm.
The Enzymatic Blockade of Protein Synthesis
Upon its successful release from the endosome, Fragment A enters the cytoplasm and is liberated from Fragment B by the reduction of the disulfide bond. Once free, the Catalytic (C) domain of Fragment A becomes an active enzyme. Its function is to sabotage the host cell’s protein-making machinery in a process known as ADP-ribosylation.
The specific target of Fragment A is Eukaryotic Elongation Factor 2 (eEF-2), a protein required for translation, the process of synthesizing new proteins. The normal function of eEF-2 is to move the growing peptide chain along the ribosome, a movement called translocation. Without active eEF-2, the ribosome stalls, and the production of all cellular proteins ceases immediately.
The biochemical reaction involves Fragment A using Nicotinamide Adenine Dinucleotide (NAD+) as a substrate. Fragment A cleaves NAD+ and transfers the resulting ADP-ribose group onto a unique amino acid residue found only on eEF-2, called diphthamide. Diphthamide is a modified histidine residue, and its presence is necessary for the toxin to act. The attachment of the ADP-ribose molecule permanently inactivates the elongation factor.
The catalytic power of diphtheria toxin is immense, as a single molecule of Fragment A can modify and inactivate thousands of eEF-2 molecules. This rapid blockade of protein synthesis swiftly leads to cell death. Because protein synthesis is fundamental to all cellular function, the toxin’s mechanism leads to the tissue necrosis and organ failure seen in severe diphtheria infections.
Utilizing the Mechanism for Immunity
Understanding the diphtheria toxin’s structure and mechanism was fundamental to developing the effective diphtheria toxoid vaccine. This vaccine relies on separating the toxin’s ability to trigger an immune response from its ability to cause cellular damage.
The vaccine is created by treating the native diphtheria toxin with a chemical agent, most commonly formalin (a solution of formaldehyde). This treatment chemically inactivates the toxin, neutralizing the enzymatic activity of Fragment A. Formaldehyde achieves this by creating cross-links and modifications within the catalytic domain, without significantly altering the protein’s overall structure.
The resulting product, called a toxoid, is non-toxic but retains the structural shape of the original toxin, especially the Fragment B domains. When administered, this toxoid stimulates the immune system to recognize the toxin’s structure and produce protective antibodies. These antibodies bind to the native toxin if infection occurs, preventing the binding and internalization steps, and stopping cellular destruction.