Batch purification of high-purity lysozyme from egg white and characterization of the enzyme modified by PEGylation.
PEGylation is one of the most promising and extensively studied strategies for improving the pharmacological properties of proteins as well as their physical and thermal stability. Purified lysozyme obtained from hen egg white by batch mode was modified by PEGylation with methoxypolyethyleneglycol succinimidyl succinato (mPEG-SS, MW 5000). The conjugates produced retained full enzyme activity with the substrate glycol chitosan, independent of degree of enzyme modification, although lysozyme activity with the substrate Micrococcus lysodeikticus was altered according to the degree of modification. The conjugate with a low degree of modification by mPEG-SS retained 67% of its enzyme activity with the M. lysodeikticus substrate. The mPEG-SS was also shown to be a highly reactive polymer. The effects of pH and temperature on PEGylated lysozymes indicated that the conjugate was active over a wide pH range and was stable up to 50 degrees C. This conjugate also showed resistance to proteolytic degradation, remained stable in human serum, and displayed greater antimicrobial activity than native lysozyme against Gram-negative bacteria.
Silva Freitas
Débora da
Dd
Department of Biochemical and Pharmaceutical Technology, Faculty of Pharmaceutical Sciences, University of São Paulo, São Paulo, Brazil. debyrp@usp.br
Abrahão-Neto
José
J
D000818:Animals; D002626:Chemistry, Pharmaceutical / Q000379:methods*; D004529:Egg White; D004789:Enzyme Activation / Q000187:drug effects / Q000502:physiology; D006801:Humans; D008837:Micrococcus / Q000187:drug effects / Q000502:physiology; D009113:Muramidase / Q000302:isolation & purification* / Q000378:metabolism / Q000494:pharmacology; D011092:Polyethylene Glycols / Q000494:pharmacology*
D011092:Polyethylene Glycols; C421864:hen egg lysozyme; D009113:Muramidase