Interfacial binding of cutinase rather than its catalytic activity determines the steady state interfacial tension during oil drop lipid hydrolysis.
Hydrolysis of triglycerides by cutinase from Fusarium solani pisi causes in oil drop tensiometer experiments a decrease of the interfacial tension. A series of cutinase variants with amino acid substitutions at its molecular surface yielded different values of the steady state interfacial tension. This tension value poorly correlated with the specific activity as such nor with the total activity (defined as the specific activity multiplied by the amount of enzyme bound) of the cutinase variants. Moreover, it appeared that at activity levels above 15% of that of wild type cutinase the contribution of hydrolysis to the decrease of the tension is saturating. A clear positive correlation was found between the interfacial tension plateau value and the interfacial binding of cutinase, as determined with attenuated total reflection Fourier transformed infrared spectroscopy (ATR-FTIR). These results indicate that the interfacial steady state level is not determined by the rate of hydrolysis, but mainly by the interfacial binding of cutinase.
Flipsen
J A
JA
Department of Enzymology and Protein Engineering, Utrecht University, The Netherlands.
van Schaick
M A
MA
Dijkman
R
R
van der Hijden
H T
HT
Verheij
H M
HM
Egmond
M R
MR
D002265:Carboxylic Ester Hydrolases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D005670:Fusarium / Q000201:enzymology / Q000235:genetics; D014644:Genetic Variation; D006868:Hydrolysis; D008055:Lipids / Q000737:chemistry; D017550:Spectroscopy, Fourier Transform Infrared; D013500:Surface Tension; D014280:Triglycerides / Q000737:chemistry
D008055:Lipids; D014280:Triglycerides; D002265:Carboxylic Ester Hydrolases; C019952:cutinase