A kinetic model for enzyme interfacial activity and stability: pa‐hydroxynitrile lyase at the diisopropyl ether/water interface

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Abstract
A kinetic framework is developed to describe enzyme activity and stability in two‐phase liquid–liquid systems. In particular, the model is applied to the enzymatic production of benzaldehyde from mandelonitrile by Prunus amygdalus hydroxynitrile lyase (pa‐Hnl) adsorbed at the diisopropyl ether (DIPE)/aqueous buffer interface (pH = 5.5). We quantitatively describe our previously obtained experimental kinetic results (Hickel et al., 1999; 2001), and we successfully account for the aqueous‐phase enzyme concentration dependence of product formation rates and the observed reaction rates at early times. Multilayer growth explains the early time reversibility of enzyme adsorption at the DIPE/buffer interface observed by both enzyme‐activity and dynamic‐interfacial‐tension washout experiments that replace the aqueous enzyme solution with a buffer solution. The postulated explanation for the unusual stability of pa‐Hnl adsorbed at the DIPE/buffer interface is attributed to a two‐layer adsorption mechanism. In the first layer, slow conformational change from the native state leads to irreversible attachment and partial loss of catalytic activity. In the second layer, pa‐Hnl is reversibly adsorbed without loss in catalytic activity. The measured catalytic activity is the combined effect of the deactivation kinetics of the first layer and of the adsorption kinetics of each layer. For the specific case of pa‐Hnl adsorbed at the DIPE/buffer interface, this combined effect is nearly constant for several hours resulting in no apparent loss of catalytic activity. Our proposed kinetic model can be extended to other interfacially active enzymes and other organic solvents. Finally, we indicate how interfacial‐tension lag times provide a powerful tool for rational solvent selection and enzyme engineering. © 2002 Wiley Periodicals, Inc. Biotechnol Bioeng 78: 595–605, 2002.
Funding Information
  • Department of Energy (DE-FG03-94ER14456)
  • Nestlé Inc., Switzerland

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