Improving the heat resistance of ribonuclease A by the addition of poly(N,N-diethylaminoethyl methacrylate)-graft-poly(ethylene glycol) (PEAMA-g-PEG).

Poly(N,N-diethylaminoethyl methacrylate)-graft-poly(ethylene glycol) (PEAMA-g-PEG) has previously been used as a novel additive to improve the heat resistance of lysozyme, which has a positive net charge and a negatively charged active site. In the present study, we show that PEAMA-g-PEG prevents heat inactivation of ribonuclease A (RNase A), which has a positive net charge and a positively charged active site. After treatment at 98 °C for 10 min, the enzymatic activity of RNase A complexed with PEAMA-g-PEG was maintained at up to 75% of the level of the native RNase A.

Indispensable structure of solution additives to prevent inactivation of lysozyme for heating and refolding.

This article investigates solution additives that prevent misfolding of lysozyme from heating treatment and during refolding processes. Comparison of heat treatment of native lysozyme and oxidative refolding from the reduced and denatured state of lysozyme in the presence of 44 different additives revealed an indispensable chemical structure for the additives to be effective against heat-induced misfolding and for refolding. The additives effective against heat treatment of native lysozyme possessed a main chain of the amino acid moiety.

Regulation of lysozyme activity based on thermotolerant protein/smart polymer complex formation.

Proteins have evolved to acquire highly specialized biological functions and are ideal for various applications in both medicine and biotechnology, although denaturation is one of the major problems in protein chemistry. Here, we show a novel strategy for the regulation and preservation of the enzymatic activity even after heat treatment by the complex formation with a cationic smart copolymer, poly(N,N-diethylaminoethyl methacrylate)-graft-poly(ethylene glycol) (PEAMA-g-PEG). PEAMA-g-PEG suppressed the enzymatic activity of lysozyme completely without any conformational change, indicating complex formation and the capping of the active site of lysozyme by PEAMA-g-PEG.

Amidated amino acids are prominent additives for preventing heat-induced aggregation of lysozyme.

An additive that is highly effective in small amounts for controlling protein inactivation and aggregation has long been demanded. In this paper we show amidated amino acids as new potent additives. In the presence of 100 mM amidated amino acids, e.