scholarly journals Single-chain immunotoxins directed at the human transferrin receptor containing Pseudomonas exotoxin A or diphtheria toxin: anti-TFR(Fv)-PE40 and DT388-anti-TFR(Fv).

1991 ◽  
Vol 11 (4) ◽  
pp. 2200-2205 ◽  
Author(s):  
J K Batra ◽  
D J Fitzgerald ◽  
V K Chaudhary ◽  
I Pastan
Keyword(s):  
Transferrin Receptor ◽  
Diphtheria Toxin ◽  
Gene Fusion ◽  
Antigen Binding ◽  
Dna Encoding ◽  
Single Chain ◽  
Pseudomonas Exotoxin ◽  
Pseudomonas Exotoxin A ◽  
Human Transferrin ◽  
Human Transferrin Receptor

Two single-chain immunotoxins directed at the human transferrin receptor have been constructed by using polymerase chain reaction-based methods. Anti-TFR(Fv)-PE40 is encoded by a gene fusion between the DNA sequence encoding the antigen-binding portion (Fv) of a monoclonal antibody directed at the human transferrin receptor and that encoding a 40,000-molecular-weight fragment of Pseudomonas exotoxin (PE40). The other fusion protein, DT388-anti-TFR(Fv), is encoded by a gene fusion between the DNA encoding a truncated form of diphtheria toxin and that encoding the antigen-binding portion of antibody to human transferrin receptor. These gene fusions were expressed in Escherichia coli, and fusion proteins were purified by conventional chromatography techniques to near homogeneity. In anti-TFR(Fv)-PE40, the antigen-binding portion is placed at the amino terminus of the toxin, while in DT388-anti-TFR(Fv), it is at the carboxyl end of the toxin. Both these single-chain immunotoxins kill cells bearing the human transferrin receptors. However, anti-TFR(Fv)-PE40 was usually more active than DT388-anti-TFR(Fv), and in some cases it was several-hundred-fold more active. Anti-TFR(Fv)-PE40 was also more active on cell lines than a conjugate made by chemically coupling the native antibody to PE40, and in some cases it was more than 100-fold more active.

Single-chain immunotoxins directed at the human transferrin receptor containing Pseudomonas exotoxin A or diphtheria toxin: anti-TFR(Fv)-PE40 and DT388-anti-TFR(Fv)

1991 ◽  
Vol 11 (4) ◽  
pp. 2200-2205
Author(s):  
J K Batra ◽  
D J Fitzgerald ◽  
V K Chaudhary ◽  
I Pastan
Keyword(s):  
Transferrin Receptor ◽  
Diphtheria Toxin ◽  
Gene Fusion ◽  
Antigen Binding ◽  
Dna Encoding ◽  
Single Chain ◽  
Pseudomonas Exotoxin ◽  
Pseudomonas Exotoxin A ◽  
Human Transferrin ◽  
Human Transferrin Receptor

Two single-chain immunotoxins directed at the human transferrin receptor have been constructed by using polymerase chain reaction-based methods. Anti-TFR(Fv)-PE40 is encoded by a gene fusion between the DNA sequence encoding the antigen-binding portion (Fv) of a monoclonal antibody directed at the human transferrin receptor and that encoding a 40,000-molecular-weight fragment of Pseudomonas exotoxin (PE40). The other fusion protein, DT388-anti-TFR(Fv), is encoded by a gene fusion between the DNA encoding a truncated form of diphtheria toxin and that encoding the antigen-binding portion of antibody to human transferrin receptor. These gene fusions were expressed in Escherichia coli, and fusion proteins were purified by conventional chromatography techniques to near homogeneity. In anti-TFR(Fv)-PE40, the antigen-binding portion is placed at the amino terminus of the toxin, while in DT388-anti-TFR(Fv), it is at the carboxyl end of the toxin. Both these single-chain immunotoxins kill cells bearing the human transferrin receptors. However, anti-TFR(Fv)-PE40 was usually more active than DT388-anti-TFR(Fv), and in some cases it was several-hundred-fold more active. Anti-TFR(Fv)-PE40 was also more active on cell lines than a conjugate made by chemically coupling the native antibody to PE40, and in some cases it was more than 100-fold more active.

scholarly journals Construction, expression and characterization of chimaeric toxins containing the ribonucleolytic toxin restrictocin: intracellular mechanism of action

1997 ◽  
Vol 324 (3) ◽  
pp. 815-822 ◽  
Author(s):  
Dharmendar RATHORE ◽  
Janendra K. BATRA
Keyword(s):  
Transferrin Receptor ◽  
Metabolic Inhibitors ◽  
Target Cells ◽  
Cytotoxic Activities ◽  
Dna Encoding ◽  
Single Chain ◽  
Human Transferrin ◽  
Human Transferrin Receptor ◽  
A Cell

Restrictocin is a ribonucleolytic toxin produced by the fungus Aspergillus restrictus. Two chimaeric toxins containing restrictocin directed at the human transferrin receptor have been constructed. Anti-TFR(scFv)–restrictocin is encoded by a gene produced by fusing the DNA encoding a single-chain antigen-combining region (scFv) of a monoclonal antibody, directed at the human transferrin receptor, at the 5′ end of that encoding restrictocin. The other chimaeric toxin, restrictocin–anti-TFR(scFv), is encoded by a gene fusion containing the DNA encoding the single-chain antigen-combining region of antibody to human transferrin receptor at the 3′ end of the DNA encoding restrictocin. These gene fusions were expressed in Escherichia coli, and fusion proteins purified from the inclusion bodies by simple chromatography techniques to near-homogeneity. The two chimaeric toxins were found to be equally active in inhibiting protein synthesis in a cell-free in vitrotranslation assay system. The chimaeric toxins were selectively toxic to the target cells in culture with potent cytotoxic activities. However, restrictocin–anti-TFR(scFv) was more active than anti-TFR(scFv)–restrictocin on all cell lines studied. By using protease and metabolic inhibitors, it can be shown that, to manifest their cytotoxic activity, the restrictocin-containing chimaeric toxins need to be proteolytically processed intracellularly and the free toxin or a fragment thereof thus generated is translocated to the target via a route involving the Golgi apparatus.

scholarly journals Structural model of phospholipid-reconstituted human transferrin receptor derived by electron microscopy

Structure ◽  
1998 ◽  
Vol 6 (10) ◽  
pp. 1235-1243 ◽  
Author(s):  
Hendrik Fuchs ◽  
Uwe Lücken ◽  
Rudolf Tauber ◽  
Andreas Engel ◽  
Reinhard Geßner
Keyword(s):  
Electron Microscopy ◽  
Transferrin Receptor ◽  
Structural Model ◽  
Human Transferrin ◽  
Human Transferrin Receptor

The Promoter Region of the Human Transferrin Receptor Gene

1988 ◽  
Vol 526 (1 Hemochromatos) ◽  
pp. 54-64 ◽  
Author(s):  
John L. Casey ◽  
Bruno Jeso ◽  
Krishnamurthy Rao ◽  
Tracey A. Rouault ◽  
Richard D. Klausner ◽  
...  
Keyword(s):  
Transferrin Receptor ◽  
Promoter Region ◽  
Receptor Gene ◽  
Human Transferrin ◽  
Human Transferrin Receptor

scholarly journals Human transferrin receptor internalization is partially dependent upon an aromatic amino acid on the cytoplasmic domain.

1990 ◽  
Vol 1 (4) ◽  
pp. 369-377 ◽  
Author(s):  
T E McGraw ◽  
F R Maxfield
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Rate Constant ◽  
Transferrin Receptor ◽  
Aromatic Amino Acid ◽  
Opposite Polarity ◽  
Receptor Internalization ◽  
Aromatic Side Chain ◽  
Human Transferrin ◽  
Human Transferrin Receptor

The objective of this work is to identify the elements of the human transferrin receptor that are involved in receptor internalization, intracellular sorting, and recycling. We have found that an aromatic side chain at position 20 on the cytoplasmic portion of the human transferrin receptor is required for efficient internalization. The wild-type human transferrin receptor has a tyrosine at this position. Replacement of the Tyr-20 with an aromatic amino acid does not alter the rate constant of internalization, whereas substitution with the nonaromatic amino acids serine, leucine, or cysteine reduces the internalization rate constant approximately three-fold. These results are consistent with similar studies of other receptor systems that have also documented the requirement for a tyrosine in rapid internalization. The amino terminus of the transferrin receptor is cytoplasmic, with the tyrosine 41 amino acids from the membrane. These two features distinguish the transferrin receptor from the other membrane proteins for which the role of tyrosine in internalization has been examined, because these proteins have the opposite polarity with respect to the membrane and because the tyrosines are located closer to the membrane (within 25 amino acids). The externalization rate for the recycling of the transferrin receptor is not altered by any of these substitutions, demonstrating that the aromatic amino acid internalization signal is not required for the efficient exocytosis of internalized receptor.

Sign in / Sign up

Export Citation Format

Share Document