Role of organic solvents on Pa-hydroxynitrile lyase interfacial activity and stability.
Catalytic activity and adsorption of Pa-hydroxynitrile lyase (Pa-Hnl) was investigated at various organic solvent/water interfaces. We focused on the role of solvent polarity in promoting activity and stability in two-phase systems, specifically for the solvents heptane, dibutyl ether (DBE), diisopropyl ether (DIPE), butylmethyl ether (BME), and methyl tert-butyl ether (MTBE). Enzyme activity towards mandelonitrile cleavage was determined in a recycle reactor with a well-defined interfacial area as described by Hickel, et al. 1999. Here the recycle reactor was modified to permit exchange of the aqueous phase. With this modification, irreversibility of enzyme adsorption was determined as a function of the adsorption time at the interface. Irreversibility of enzyme adsorption was also investigated by measuring the surface pressure of a sessile-drop upon washout. We find that Pa-Hnl exhibits the highest stability but the lowest initial catalytic activity at the aqueous/organic solvent interface with the most polar organic solvents. Thus, DIPE and MTBE display no loss in enzyme activity over a period of several hours. However, the more apolar the solvent is the higher the initial Pa-Hnl activity, but the faster the loss of activity. Dynamic tensiometry reveals that Pa-Hnl adsorbs more strongly at the interface of the more apolar solvents. Surprisingly, Pa-Hnl develops some irreversible adsorption after 30 min at the DIPE/water interface, but does not lose catalytic activity.
Hickel
A
A
Institut für Biophysik und Röntgenstrukturforschung, Schmiedlstrasse 6, 8042 Graz, Austria. andrea.hickel@oeaw.ac.at
Radke
C J
CJ
Blanch
H W
HW
D000327:Adsorption; D000446:Aldehyde-Lyases / Q000378:metabolism*; D002384:Catalysis; D004795:Enzyme Stability; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D009754:Nuts; D011489:Protein Denaturation; D019685:Rosales / Q000201:enzymology; D012997:Solvents / Q000378:metabolism*; D014867:Water
D012997:Solvents; D014867:Water; D000446:Aldehyde-Lyases; C079359:hydroxymandelonitrile lyase