Hi. I´m trying to immobilize a lipase in a polypropylene support. Please, is there a process to increase the contact between this carrier and the enzyme?, because due the hydrophobicity of the polypropilene, it is always located on the surface of the buffer, despite the continuos stirring. I have not not get immobilize the enzyme ( the activity of the buffer media after 48h, does not changed through the experiment). Previously I washed the carrier with an ethanol solution (50%) and rinsed with water. Thanks so much in advance for any help or suggestion.
Use gluteraldehyde or formaldehyde (1-5%) as cross linker. they undergo Schiff base reaction (CHO) of gluteraldehyde combine with NH2 group of protein). This reaction is only stable in alkaline condition, While at acidic condition use any stablizer to stable Schiff base reaction.
Add substrate or an additive as described in
https://www.researchgate.net/publication/255735294_Improving_Lipase_Activity_by_Immobilization_and_Post-immobilization_Strategies
Article Improving Lipase Activity by Immobilization and Post-immobil...
Hi Ingrid. The fact that you didn't observe a decrease of enzyme activity in the contactning buffer solution may be due to low specific surface area of the polypropylene (PP) powder/beads. I guess your immobilization by adsorption is intended for catalysis in organic medium (for use in aqueous medium you would need a covalent binding). For the use in organics it may be enough to precipitate/spread the enzyme on the powder surface during evaporation of the enzyme solution-PP mixture in rotary evaporator. For better wettability of PP powder you may use its treatment with ethanol, which you mentioned, followed by transfer into the buffer solution and vacuumization to get rid of air bubbles. This may decrease the floating of the powder. I succeeded to wetten 50 -100 mkm polystyrene beads in this way.
Best wishes
Alexander
Thanks doctor for your kind response and help. I'm going to evaluate the procedure with rotary evaporator just you recommended. The treatment with ethanol, buffer washing and vacuum I already done but the powder remains on the surface of the enzyme solution. Do you recommend a specif concentration of ethanol per gram of pp? . I reviewed some publications and there are different approaches. Thanks again for your help
Onto the hydrophobic support it is better to immobilize the enzyme by Physical adsorption. But your protein loading might be less by Physical adsorption. You may apply some cross linker after physical adsorption to achieve a stable biocatalyst.
Best wishes
Use gluteraldehyde or formaldehyde (1-5%) as cross linker. they undergo Schiff base reaction (CHO) of gluteraldehyde combine with NH2 group of protein). This reaction is only stable in alkaline condition, While at acidic condition use any stablizer to stable Schiff base reaction.
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