Constructing a DNAzyme Delivery and Expression System for Escherichia coli: A Prototype for Phage Therapy

Abstract

Antisense oligonucleotides exhibit high potential for use as therapeutic agents. '10-23' DNAzymes are antisense molecules with a high chemical stability and catalytic efficiency. In the present study, we developed a phagemid containing a DNAzyme expression system regulated by two promoters. One of these promoters, pA1, promotes constitutive expression of Moloney murine leukemia virus reverse transcriptase (MoMuLV-RT). The other promoter, plac, regulates transcription of the RNA substrate from which MoMuLV-RT produces the DNAzyme by reverse transcription. The ftsZ DNAzyme was used to validate this expression system in the phagemid, named pDESCP. ftsZ DNAzyme expression altered the morphological pattern of Escherichia coli from a bacillary to filamentous form. In E. coli FtsZ is the primary component of the cell division apparatus, forming a structure known as Z-ring, which is the place of division. It is suggested that the DNAzyme ftsZ is decreasing the translation of this protein. Delivery of pDESCP into F+ strain of E. coli cells, using VCSM13, and the possible insertion of other DNAzymes into the cassette makes this phagemid an important prototype for phage therapy.