Functional and structural characterization of chimeras of a bacterial genotoxin and human type I DNAse

Abstract

Chimeras composed of the cdtB gene of a novel bacterial genotoxin and the human type I deoxyribonuclease (DNase I) gene were constructed and their products characterized relative to the biochemical and enzymatic properties of the native proteins. The product of a cdtB/DNase I chimera formed a heterotrimer with the CdtA and CdtC subunits of the genotoxin and targeted mutations increased the specific activity of the hybrid protein. Expression of active chimeric gene products established that the CdtB protein is an atypical divalent cation-dependent endonuclease and demonstrated the potential for genetically engineering a new class of therapeutic agent for inhibiting the proliferation of cancer cells.

Cytolethal Distending Toxin Damages the Oral Epithelium of Gingival Explants

“…Mutated subunit CdtB H160A forms a heterotrimer with wild-type CdtA and CdtC that fails to inhibit the proliferation of susceptible cells (DiRienzo et al, 2009). Tissue exposed to 10 μg/mL of toxin reconstituted with CdtAB H160A C appeared histologically similar to untreated tissue (Fig.…”

A New Functional Site W115 in CdtA Is Critical for Aggregatibacter actinomycetemcomitans Cytolethal Distending Toxin

“…There is clear evidence that CdtB functions as a type I deoxyribonuclease (DNase I), whereby it activates a DNA-damage-dependent checkpoint that eventually leads cells to apoptosis [13]–[15]. Mutations of the DNase I active site residues in CdtB resulted in the loss of nuclease ability and cell cycle arrest in vivo [16], [17].…”

Localization of Aggregatibacter actinomycetemcomitans Cytolethal Distending Toxin Subunits during Intoxication of Live Cells

“…The toxic third subunit (CdtB) potentially has multiple activities based on its phylogenetic relationship to a superfamily of enzymes that includes the endonucleases, exonucleases, sphingomyelinases, and inositol polyphosphate 5-phosphatases (15). However, the general consensus is that the primary in vivo action of the Cdt is that of a genotoxin (8,9,19,23,32). The result of intoxication with the Cdt is cell cycle arrest, leading to the more definitive classification of this group of toxins as cyclomodulins (31,33).…”