Thermal aggregation of alpha-chymotrypsin: role of hydrophobic and electrostatic interactions.
We have recently reported that electrostatic interactions may play a critical role in alcohol-induced aggregation of alpha-chymotrypsin (CT). In the present study, we have investigated the heat-induced aggregation of this protein. Thermal aggregation of CT obeyed a characteristic pattern, with a clear lag phase followed by a sharp rise in turbidity. Intrinsic and ANS fluorescence studies, together with fluorescence quenching by acrylamide, suggested that the hydrophobic patches are more exposed in the denatured conformation. Typical chaperone-like proteins, including alpha- and beta-caseins and alpha-crystalline could inhibit thermal aggregation of CT, and their inhibitory effect was nearly pH-independent (within the pH range of 7-9). This was partially counteracted by alpha-, beta- and especially gamma-cyclodextrins, suggesting that hydrophobic interactions may play a major role. Loss of thermal aggregation at extreme acidic and basic conditions, combined with changes in net charge/pH profile of aggregation upon chemical modification of lysine residues are taken to support concomitant involvement of electrostatic interactions.
Rezaei-Ghaleh
Nasrollah
N
Institute of Biochemistry and Biophysics, University of Tehran, Tehran, Iran.
Ramshini
Hassan
H
Ebrahim-Habibi
Azadeh
A
Moosavi-Movahedi
Ali Akbar
AA
Nemat-Gorgani
Mohsen
M
D002364:Caseins / Q000737:chemistry; D002918:Chymotrypsin / Q000037:antagonists & inhibitors / Q000737:chemistry*; D004795:Enzyme Stability; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D057927:Hydrophobic and Hydrophilic Interactions; D018832:Molecular Chaperones / Q000737:chemistry; D011487:Protein Conformation; D011489:Protein Denaturation; D055672:Static Electricity; D038201:alpha-Crystallins / Q000737:chemistry
D002364:Caseins; D018832:Molecular Chaperones; D038201:alpha-Crystallins; D002918:Chymotrypsin; C117608:alpha-chymotrypsin