Mutagenesis of the Clostridium difficile toxin B gene and effect on cytotoxic activity
| Autor: | David M. Lyerly, J. Scott Moncrief, Lisa Barroso, Tracy D. Wilkins |
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| Rok vydání: | 1994 |
| Předmět: |
Mutant
Bacterial Toxins Molecular Sequence Data Clostridium difficile toxin B CHO Cells Biology Microbiology Serine Enterotoxins Bacterial Proteins Cricetinae Escherichia coli Animals Nucleotide Histidine Amino Acid Sequence Cysteine Binding site Site-directed mutagenesis Sequence Deletion chemistry.chemical_classification Clostridioides difficile Mutagenesis Molecular biology Infectious Diseases Biochemistry chemistry Genes Bacterial Mutagenesis Site-Directed |
| Zdroj: | Microbial pathogenesis. 16(4) |
| ISSN: | 0882-4010 |
| Popis: | Toxins A and B of Clostridium difficile are large cytotoxic proteins that share several unusual structural features, including four conserved cysteines, a potential nucleotide binding site, a hydrophobic region, and a series of contiguous repeating units at the carboxyl terminus. In the following study, we developed a series of toxin B mutants with altered properties in each of these features and examined the effect of the mutation on cytotoxic activity. Altering conserved cysteines to serine resulted in a 90% reduction in activity, whereas altering a histidine residue located in the potential nucleotide binding site to glutamine resulted in a 99% reduction. Removing the repeating units lowered the activity by 90% whereas removing the repeating units plus a conserved cysteine located just upstream of the units reduced the activity by more than five logs, resulting in an inactive toxin. Deleting the internal hydrophobic region had a similar effect. Our findings demonstrate that these conserved features appear to be important for expression of cytotoxic activity. |
| Databáze: | OpenAIRE |
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