Bacterial biofilm degradation by recombinant SpdAZ cloned from Streptococcus pyogenes ADUYE1.

Nucleases break down nucleic acids into smaller pieces or monomers. These enzymes are important in many biological activities, such as obtaining nucleotides necessary for cell division, DNA repair and recombination, fragmenting DNA during apoptosis, as well as functioning as an infectious agent or contributing to host defense mechanisms and disrupting bacterial biofilm structures. Herein, a nuclease from Streptococcus pyogenes (S. pyogenes) ADUYE1, homologous to the spd3 gene and named as spdAZ, was cloned and heterologously expressed in Escherichia coli (E. coli). Total protein was extracted from transformed E. coli and recombinant SpdAZ (rSpdAZ) was purified using IMAC method. Sequencing analysis of the cloned gene showed 5 amino acid substitutions between Spd3 and SpdAZ. The DNAse activity of the purified rSpdAZ was tested on viral, bacterial and eukaryotic DNA as well as with DNase agar. The anti-biofilm activity of rSpdAZ was tested against biofilms formed by 8 bacterial isolates, including Pseudomonas aeruginosa, E. coli, methicillin-resistant Staphylococcus aureus (MRSA), and methicillin-resistant Staphylococcus epidermidis (MRSE). Our results showed that rSpdAZ exhibited nuclease activity on all the DNA samples tested. rSpdAZ enzyme was effective against the biofilms formed by all the tested bacteria. While the effect of rSpdAZ in reducing the mature biofilm layers ranged between 65 % and 93 %, its effect in preventing biofilm formation (i.e., pre-biofilm) was between 48 % and 91 %. Enzyme activity against mature biofilms occurred after 4 h in all studied species Biofilm formation is one of the main problems to fight against bacteria by decreasing efficacy of the antibacterial agents used. Our data suggest that rSpdAZ may be used as an antibiofilm agent.