Mechanism of solvent induced thermal stabilization of papain.
In the present study an attempt is made to elucidate the effects of various cosolvents, such as sorbitol, sucrose, xylose and glycerol, on papain. The stabilizing effects of these cosolvents on the structure and function of papain is determined by the activity measurements, fluorescence spectroscopy and differential scanning calorimetry (DSC). The enzyme activity measurements indicate several fold increase in the thermal stability of the enzyme in all the cosolvents used. The thermal denaturation studies of papain in presence of various concentrations of cosolvents indicated a shift in the apparent thermal denaturation temperature (app Tm) suggesting increased thermal stability of papain in presence of cosolvents. The app Tm shifted from a control value of 83+/-1 degrees C to a value of >90+/-1 degrees C in presence of 40% sorbitol. The DSC thermogram for native papain can be clearly deconvoluted into two transitions corresponding to left and right domain and in presence of cosolvents both transitions A and B shift to higher temperature. Maximum stabilization was seen in case of 30% sorbitol where the thermal transition temperatures increased compared to control. The results from partial specific volume measurements of papain in presence of cosolvents suggest that the preferential interaction parameter (xi3) was negative in all cosolvents and maximum hydration was observed in the case of glycerol where the preferential interaction parameter was 0.165g/g. These above results suggest that there is a considerable increase in the thermal stability of papain in presence of these cosolvents as a result of preferential hydration.
Sathish
H A
HA
Department of Protein Chemistry and Technology, Central Food Technological Research Institute, Mysore 570 020, India.
Kumar
Parigi Ramesh
PR
Prakash
V
V
D002152:Calorimetry, Differential Scanning; D004305:Dose-Response Relationship, Drug; D004795:Enzyme Stability / Q000187:drug effects*; D006358:Hot Temperature; D010206:Papain / Q000737:chemistry / Q000378:metabolism*; D011489:Protein Denaturation; D012997:Solvents / Q000737:chemistry* / Q000494:pharmacology*; D013050:Spectrometry, Fluorescence
D012997:Solvents; D010206:Papain