Understanding structure-stability relationships of Candida antartica lipase B in ionic liquids.
Two different water-immiscible ionic liquids (ILs), 1-ethyl-3-methylimidizolium bis(trifluoromethylsulfonyl)imide and butyltrimethylammonium bis(trifluoromethylsulfonyl)imide, were used for butyl butyrate synthesis from vinyl butyrate catalyzed by Candida antarctica lipase B (CALB) at 2% (v/v) water content and 50 degrees C. Both the synthetic activity and stability of the enzyme in these ILs were enhanced as compared to those in hexane. Circular dichroism and intrinsic fluorescence spectroscopic techniques have been used over a period of 4 days to determine structural changes in the enzyme associated with differences in its stability for each assayed medium. CALB showed a loss in residual activity higher than 75% after 4 days of incubation in both water and hexane media at 50 degrees C, being related to great changes in both alpha-helix and beta-strand secondary structures. The stabilization of CALB, which was observed in the two ILs studied, was associated with both the maintenance of the 50% of initial alpha-helix content and the enhancement of beta-strands. Furthermore, intrinsic fluorescence studies clearly showed how a classical enzyme unfolding was occurring with time in both water and hexane media. However, the structural changes associated with the incubation of the enzyme in both ILs might be attributed to a compact and active enzyme conformation, resulting in an enhancement of the stability in these nonaqueous environments.
De Diego
Teresa
T
Departamento de Bioquímica y Biología Molecular B e Inmunología, Facultad de Química, Universidad de Murcia, P.O. Box 4021, E-30100 Murcia, Spain.
Lozano
Pedro
P
Gmouh
Said
S
Vaultier
Michel
M
Iborra
José L
JL
D000595:Amino Acid Sequence / Q000235:genetics; D004795:Enzyme Stability / Q000235:genetics; D005656:Fungal Proteins; D008049:Lipase / Q000737:chemistry* / Q000235:genetics*; D008969:Molecular Sequence Data; D009994:Osmolar Concentration; D017433:Protein Structure, Secondary / Q000235:genetics; D012996:Solutions / Q000737:chemistry*
D005656:Fungal Proteins; D012996:Solutions; D008049:Lipase; C431349:lipase B, Candida antarctica