Why is trehalose an exceptional protein stabilizer? An analysis of the thermal stability of proteins in the presence of the compatible osmolyte trehalose.
Trehalose, a naturally occurring osmolyte, is known to be an exceptional stabilizer of proteins and helps retain the activity of enzymes in solution as well as in the freeze-dried state. To understand the mechanism of action of trehalose in detail, we have conducted a thorough investigation of its effect on the thermal stability in aqueous solutions of five well characterized proteins differing in their various physico-chemical properties. Among them, RNase A has been used as a model enzyme to investigate the effect of trehalose on the retention of enzymatic activity upon incubation at high temperatures. 2 m trehalose was observed to raise the transition temperature, Tm of RNase A by as much as 18 degrees C and Gibbs free energy by 4.8 kcal mol-1 at pH 2.5. There is a decrease in the heat capacity of protein denaturation (DeltaCp) in trehalose solutions for all the studied proteins. An increase in the DeltaG and a decrease in the DeltaCp values for all the proteins points toward a general mechanism of stabilization due to the elevation and broadening of the stability curve (DeltaG versus T). A direct correlation of the surface tension of trehalose solutions and the thermal stability of various proteins has been observed. Wyman linkage analysis indicates that at 1.5 m concentration 4-7 molecules of trehalose are excluded from the vicinity of protein molecules upon denaturation. We further show that an increase in the stability of proteins in the presence of trehalose depends upon the length of the polypeptide chain. The pH dependence data suggest that even though the charge status of a protein contributes significantly, trehalose can be expected to work as a universal stabilizer of protein conformation due to its exceptional effect on the structure and properties of solvent water compared with other sugars and polyols.
Kaushik
Jai K
JK
Centre for Biotechnology, Jawaharlal Nehru University, New Delhi 110067, India.
Bhat
Rajiv
R
D000818:Animals; D002417:Cattle; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D002919:Chymotrypsinogen / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism; D003574:Cytochrome c Group / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism; D004355:Drug Stability; D004795:Enzyme Stability / Q000187:drug effects; D005079:Excipients / Q000494:pharmacology*; D066298:In Vitro Techniques; D007700:Kinetics; D009113:Muramidase / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism; D010940:Plant Proteins / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism; D011489:Protein Denaturation / Q000187:drug effects; D011506:Proteins / Q000737:chemistry* / Q000187:drug effects* / Q000378:metabolism; D012259:Ribonuclease, Pancreatic / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism; D012996:Solutions; D013500:Surface Tension; D013696:Temperature; D013816:Thermodynamics; D014199:Trehalose / Q000494:pharmacology*; D014361:Trypsin Inhibitors; D000516:alpha-Amylases / Q000037:antagonists & inhibitors
D003574:Cytochrome c Group; D005079:Excipients; D010940:Plant Proteins; D011506:Proteins; D012996:Solutions; D014361:Trypsin Inhibitors; D002919:Chymotrypsinogen; D014199:Trehalose; D012259:Ribonuclease, Pancreatic; D000516:alpha-Amylases; D009113:Muramidase