Back to Search
Start Over
Receptor-directed chimeric toxins created by sortase-mediated protein fusion.
- Source :
-
Molecular cancer therapeutics [Mol Cancer Ther] 2013 Oct; Vol. 12 (10), pp. 2273-81. Date of Electronic Publication: 2013 Aug 14. - Publication Year :
- 2013
-
Abstract
- Chimeric protein toxins that act selectively on cells expressing a designated receptor may serve as investigational probes and/or antitumor agents. Here, we report use of the enzyme sortase A (SrtA) to create four chimeric toxins designed to selectively kill cells bearing the tumor marker HER2. We first expressed and purified: (i) a receptor recognition-deficient form of diphtheria toxin that lacks its receptor-binding domain and (ii) a mutated, receptor-binding-deficient form of anthrax-protective antigen. Both proteins carried at the C terminus the sortase recognition sequence LPETGG and a H₆ affinity tag. Each toxin protein was mixed with SrtA plus either of two HER2-recognition proteins--a single-chain antibody fragment or an Affibody--both carrying an N-terminal G₅ tag. With wild-type SrtA, the fusion reaction between the toxin and receptor-recognition proteins approached completion only after several hours, whereas with an evolved form of the enzyme, SrtA*, the reaction was virtually complete within 5 minutes. The four fusion toxins were purified and shown to kill HER2-positive cells in culture with high specificity. Sortase-mediated ligation of binary combinations of diverse natively folded proteins offers a facile way to produce large sets of chimeric proteins for research and medicine.<br /> (©2013 AACR.)
- Subjects :
- Amino Acid Sequence
Antigens, Bacterial therapeutic use
Bacterial Toxins therapeutic use
Binding Sites
Cell Line, Tumor
Diphtheria Toxin therapeutic use
Gene Expression Regulation, Neoplastic drug effects
Humans
Immunotoxins therapeutic use
Neoplasms drug therapy
Neoplasms genetics
Neoplasms pathology
Protein Binding
Receptor, ErbB-2 genetics
Recombinant Fusion Proteins therapeutic use
Aminoacyltransferases genetics
Antigens, Bacterial genetics
Bacterial Proteins genetics
Bacterial Toxins genetics
Cysteine Endopeptidases genetics
Diphtheria Toxin genetics
Immunotoxins genetics
Recombinant Fusion Proteins genetics
Subjects
Details
- Language :
- English
- ISSN :
- 1538-8514
- Volume :
- 12
- Issue :
- 10
- Database :
- MEDLINE
- Journal :
- Molecular cancer therapeutics
- Publication Type :
- Academic Journal
- Accession number :
- 23945077
- Full Text :
- https://doi.org/10.1158/1535-7163.MCT-13-0358