Thermal treatment of the bacteriophage lysate of Klebsiella pneumoniae B5055 as a step for the purification of capsular depolymerase enzyme.
The lytic bacteriophage which produces the hydrolase enzyme capable of depolymerizing exopolysaccharide of Klebsiella pneumoniae and of other bacteria was isolated earlier. In this study, a simple method of depolymerase purification from the phage lysate by dissociating the enzyme from the phage particle was developed. The bacteriophage showed a relatively smaller plaque size surrounded by a wide halo indicating a depolymerase action on the capsular polysaccharide of K. pneumoniae B5055. The depolymerase activity was associated predominantly with the phage particles. Different methods have been used by various researchers to dissociate the enzyme associated with phage particles either by exposure to chemicals or by altering the environmental conditions. In this study, the potential application of thermal treatment of the bacteriophage lysate was evaluated as a step for the purification of depolymerase in comparison to the mild acid treatment method of Rieger et al. (1975). The results showed that the relative thermal stability of phage depolymerase at 60°C for 30min was the basis for harvesting the enzyme leading to disintegration of all phage particles in the lysate. Both thermal and mild acid treatment resulted in comparable enzyme levels, however; mild acid treatment appeared to be cumbersome and cause chemical contamination.
Department of Microbiology, Basic Medical Sciences (BMS) Block, Panjab University, Chandigarh 160014, India.
Tesfaye
T
Kassa
Sanjay
S
Chhibber
D001435:Bacteriophages / Q000201:enzymology*; D006026:Glycoside Hydrolases / Q000302:isolation & purification*; D006358:Hot Temperature; D007711:Klebsiella pneumoniae / Q000821:virology*; D013048:Specimen Handling / Q000379:methods; D014764:Viral Proteins / Q000302:isolation & purification*
D014764:Viral Proteins; D006026:Glycoside Hydrolases