Haloalkane hydrolysis with an immobilized haloalkane dehalogenase.
Haloalkane dehalogenase from Rhodococcus rhodochrous was covalently immobilized onto a polyethyleneimine impregnated gamma-alumina support. The dehalogenating enzyme was found to retain greater than 40% of its original activity after immobilization, displaying an optimal loading (max. activity/supported protein) of 70 to 75 mg/g with an apparent maximum (max. protein/support) of 156 mg/g. The substrate, 1,2,3-trichloropropane, was found to favorably partition (adsorb) onto the inorganic alumina carrier (10 to 20 mg/g), thereby increasing the local reactant concentration with respect to the catalyst's environment, whereas the product, 2,3-dichloropropan-1-ol, demonstrated no affinity. Additionally, the inorganic alumina support exhibited no adverse effects because of solvent/component incompatibilities or deterioration due to pH variance (pH 7.0 to 10.5). As a result of the large surface area to volume ratio of the support matrix and the accessibility of the bound protein, the immobilized biocatalyst was not subject to internal mass transfer limitations. External diffusional restrictions could be eliminated with simple agitation (mixing speed: 50 rpm; flux: 4.22 cm/min). The pH-dependence of the immobilized dehalogenase was essentially the same as that for the native enzyme. Finally, both the thermostability and resistance toward inactivation by organic solvent were improved by more than an order of magnitude after immobilization.
Dravis
B C
BC
Department of Chemical and Petroleum Engineering, 1249 Benedum Hall, University of Pittsburgh, Pittsburgh, PA 15261, USA.
Swanson
P E
PE
Russell
A J
AJ
D000537:Aluminum Oxide / Q000378:metabolism; D002384:Catalysis; D020099:Coated Materials, Biocompatible; D004795:Enzyme Stability / Q000187:drug effects / Q000502:physiology; D004800:Enzymes, Immobilized / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D006867:Hydrolases / Q000378:metabolism*; D007700:Kinetics; D008433:Mathematics; D008855:Microscopy, Electron, Scanning; D011094:Polyethyleneimine / Q000378:metabolism; D011407:Propane / Q000031:analogs & derivatives* / Q000493:pharmacokinetics; D012240:Rhodococcus / Q000201:enzymology*
D020099:Coated Materials, Biocompatible; D004800:Enzymes, Immobilized; D011094:Polyethyleneimine; D006867:Hydrolases; C046137:haloalkane dehalogenase; D000537:Aluminum Oxide; D011407:Propane