Immobilization of active alpha-chymotrypsin on RF-plasma-functionalized polymer surfaces

Various polymeric surfaces (polyester, polyethylene, polystyrene) were func tionalized under oxygen and dichlorosilane-RF-coId-plasma environments and were employed as substrates for further in situ derivatization reactions an d immobilization of alpha-Chymotrypsin. The nature and morphology of the de rivatized substrates and the substrates with immobilized enzymes were analy zed using survey and high-resolution X-ray photoelectron spectroscopy, atte nuated total reflectance-fourier transform infrared (ATR-FTIR), laser desor ption fourier transform ion cyclotron resonance mass spectrometry, chemical derivatization, and atomic force microscopy (AFM) techniques. It was demon strated that the tacticity of the polystyrene substrate did not notably inf luence the activity of the immobilized enzyme, however, spacer molecules in tercalated between the polymeric substrates (e.g., polyethylene) and the en zyme significantly increased the enzyme activity (comparable with that of t he free enzyme). Computer-aided conformational modeling of the substrate-sp acer systems indicated that the longer the spacer chain, the greater the mo bility of the enzyme. It is suggested that the greater mobility of the enzy me molecules is responsible for the enhanced activity. It has also been sho wn that the stability of the immobilized enzyme systems was good; they reta ined their activity during several washing/assay cycles. (C) 2000 John Wile y & Sons, Inc.