Activity and Adsorption of Lipase fromHumicolalanuginosaon Surfaces with Different Wettabilities

Abstract

The adsorption of Humicola lanuginosa lipase and the activity of the adsorbed lipase were studied as a function of surface wettability. The adsorption was measured by in situ ellipsometry, and the surfaces used were methylated silica surfaces. The activity of the adsorbed lipase was measured after rinsing of the cuvette, i.e., with no lipase in the bulk solution, in situ by the hydrolysis of p-nitrophenyl acetate. The lipase adsorption and activity measurements were made at concentrations of the lipase in the range 63−1050 nM, and from the surface concentrations the specific activity was calculated. The study was carried out using fully methylated surfaces (hydrophobic; 90° water contact angle) and surfaces with a higher wettability (80°, 75°, and 62° water contact angle). All experiments were performed in 3-(N-morpholino)propanesulphonic acid (MOPS) buffer at pH 7.5. The adsorbed amount was found to be highest at lipase concentrations in the range 200−300 nM and decreased with increasing wettability of the surfaces. The fraction desorbable upon dilution of the adsorbed lipase was found to decrease with the concentration. The specific activity of the lipase was found to increase with increasing wettability of the surface, probably due to changes in the orientation/conformation of the adsorbed lipase. Additional experiments were performed where lipase was adorbed to surfaces with a water contact angle of 90°, in the presence of 0.03 M CaCl2. The lipase concentrations were then 63 and 1050 nM. The amount adsorbed was significantly increased in the presence of CaCl2, but no value for the specific activity of the adsorbed lipase could be calculated due to the high activity originating from lipase adsorbed to the background surfaces (cuvette walls, Teflon tubing, and magnetic stirrer).

Production of margarine fats by enzymatic interesterification with silica‐granulated Thermomyces lanuginosa lipase in a large‐scale study

“…The optimal reaction temperature for free T. langinosa lipase is around 35-40°C in a solvent system (10). After silicagranulation, Lipozyme TL IM was relatively stable in the solvent-free system in the range from 55 to 80°C (Fig.…”

“…Lipozyme TL IM has sn-1,3 specificity and hydrophilic character. It is less expensive than the commonly applied commercial lipase Lipozyme IM (Rhizomucor miehei lipase) and offers an opportunity for industry to reduce the process cost and make the production of low-price plastic fats economically competitive to the conventional chemical randomization.Previously T. lanuginosa lipase was investigated mostly in solvent systems in free-state or immobilized forms (2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14). From kinetic studies, it was found that immobilized T. lanuginosa lipase had different characteristics depending on the solvent systems and the hydrophobicity of carriers (2,7).…”

“…Previously T. lanuginosa lipase was investigated mostly in solvent systems in free-state or immobilized forms (2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14). From kinetic studies, it was found that immobilized T. lanuginosa lipase had different characteristics depending on the solvent systems and the hydrophobicity of carriers (2,7).…”

Electrostatic‐induced interfacial assembly of enzymes with nanosheets: Controlled orientation and optimized activity

“…5,6 A part of active sites might thus be buried and inaccessible, leading to a dramatic decrease in the bioactivity compared with the native enzymes. Few attempts to control the enzymatic orientation through tailoring the wetting properties 7 or surface charges 8 of the supports revealed that the rational artificial interference was not infeasible. However, in the approaches already reported, a reactive terminal tag or a protein conjugate had to be used.…”

“…However, in the approaches already reported, a reactive terminal tag or a protein conjugate had to be used. 7,8 Recently, efforts have been made toward the orientated attachment of proteins to solid surfaces through electrostatic assembly. [8][9][10] The electrostatic recognition strategy is thus believed to be a simple and biocompatible alternative to control the protein orientation.…”