Quantitative analysis of the thermal stability of the gamma phage endolysin PlyG: A biophysical and kinetic approach to assaying therapeutic potential

• Published in: Virology (New York, N.Y.) N.Y.), 2015-03, Vol.477, p.125-132
• Main Authors: Heselpoth, Ryan D, Owens, Jacqueline M, Nelson, Daniel C
• Format: Article
• Language: English
• Subjects: Anthrax; Anti-Bacterial Agents - chemistry; Anti-Bacterial Agents - radiation effects; Antimicrobial; Bacillus anthracis - enzymology; Bacteriophage; Endolysin; Enzyme Stability; Infectious Disease; Kinetics; N-Acetylmuramoyl-L-alanine Amidase - chemistry; N-Acetylmuramoyl-L-alanine Amidase - radiation effects; PlyG; Protein Denaturation - radiation effects; Protein Stability; Temperature; Thermal stability; Viral Proteins - chemistry; Viral Proteins - radiation effects
• ISSN: 0042-6822; 1096-0341
• DOI: 10.1016/j.virol.2014.11.003

Abstract Endolysins are lytic enzymes encoded by bacteriophage that represent an emerging class of protein therapeutics. Considering macromolecular thermoresistance correlates with shelf life, PlyG, a Bacillus anthracis endolysin, was thermally characterized to further evaluate its therapeutic potential. Results from a biophysical thermal analysis revealed full-length PlyG and its isolated domains comprised thermal denaturation temperatures exceeding 63 °C. In the absence of reducing agent, PlyG was determined to be kinetically unstable, a finding hypothesized to be attributable to the chemical oxidation of cysteine and/or methionine residues. The presence of reducing agent kinetically stabilized the endolysin, with PlyG retaining at least ~50% residual lytic activity after being heated at temperatures up to 80 °C and remaining enzymatically functional after being boiled. Furthermore, the endolysin had a kinetic half-life at 50 °C and 55 °C of 35 and 5.5 h, respectively. PlyG represents a thermostable proteinaceous antibacterial with subsequent prolonged therapeutic shelf life expectancy.