Fluorescence and CD spectroscopic analysis of the alpha-chymotrypsin stabilization by the ionic liquid, 1-ethyl-3-methylimidazolium bis[(trifluoromethyl)sulfonyl]amide.
The stability of alpha-chymotrypsin in the ionic liquid, 1-ethyl-3-methyl-imidizolium bis[(trifluoromethyl)sulfonyl]amide ([emim][NTf2]), was studied at 30 and 50 degrees C and compared with the stability in other liquid media, such as water, 3 M sorbitol, and 1-propanol. The kinetic analysis of the enzyme stability pointed to the clear denaturative effect of 1-propanol, while both 3M sorbitol and [emim][NTf2] displayed a strong stabilizing power. For the first time, it is shown that enzyme stabilization by ionic liquids seems to be related to the associated structural changes of the protein that can be observed by differential scanning calorimetry (DSC) and fluorescence and circular dichroism (CD). The [emim][NTf2] enhanced both the melting temperature and heat capacity of the enzyme compared to the other media assayed. The fluorescence spectra clearly showed the ability of [emim][NTf2] to compact the native structural conformation of alpha-chymotrypsin, preventing the usual thermal unfolding which occurs in other media. Changes in the secondary structure of this beta/beta protein, as quantified by the CD spectra, pointed to the great enhancement (up 40% with respect to that in water) of beta-strands in the presence of the ionic liquid, which reflects its stabilization power.
De Diego
Teresa
T
Departamento de Bioquímica y Biología Molecular B e Inmunología, Facultad de Química, Universidad de Murcia, PO Box 4021, E-30100 Murcia, Spain.
Lozano
Pedro
P
Gmouh
Said
S
Vaultier
Michel
M
Iborra
José L
JL
D002918:Chymotrypsin / Q000032:analysis* / Q000737:chemistry*; D002942:Circular Dichroism / Q000379:methods*; D004789:Enzyme Activation; D004795:Enzyme Stability; D007093:Imidazoles / Q000737:chemistry*; D007477:Ions; D007700:Kinetics; D011487:Protein Conformation; D011489:Protein Denaturation; D017433:Protein Structure, Secondary; D012996:Solutions; D012997:Solvents / Q000737:chemistry*; D013050:Spectrometry, Fluorescence / Q000379:methods*; D013449:Sulfonamides / Q000737:chemistry*; D013696:Temperature
C500270:1-ethyl-3-methyl-imidizolium bis((trifluoromethyl)sulfonyl)amide; D007093:Imidazoles; D007477:Ions; D012996:Solutions; D012997:Solvents; D013449:Sulfonamides; D002918:Chymotrypsin; C117608:alpha-chymotrypsin