His-tag presence modulates enzymatic activity both in solution and at lipid interfaces.

FLORES, SANDRA; PERILLO, MARÍA A.; NOLAN, VERONICA; SANCHEZ, JULIETA M

Buenos Aires

Congreso; Reunion Anual SAB; 2021

Sociedad Argentina de biofisica

β-Galactosidase (β-Gal) is an important biotechnological enzyme used in the dairyindustry, pharmacology and in molecular biology. This enzyme has a commercialapplication for lactose hydrolysis in dairy products. Milk processing with β-Gal before milkis commercialized is important to solve nutritional (lactose intolerance) and technological(crystallization of dairy products) problems. In this context, it is important that the activityof β-Gal be evaluated in heterogeneous media.β-Galactosidase (β-Gal) is an importantbiotechnological enzyme used in the dairy industry, pharmacology and in molecularbiology. This enzyme has a commercial application for lactose hydrolysis in dairy products.Milk processing with β-Gal before milk is commercialized is important to solve nutritional(lactose intolerance) and technological (crystallization of dairy products) problems. In thiscontext, it is important that the activity of β-Gal be evaluated in heterogeneous media.In our laboratory we have overexpressed a recombinant β-galactosidase in Escherichia coli(E. coli). This enzyme differs from its native version (β-GalWT) in that 6 histidine residueshave been added to the carboxyl terminus in the primary sequence (β-GalHis), whichallows its purification by immobilized metal affinity chromatography (IMAC). In this workwe compared the functionality of both proteins and evaluated their catalytic behavior onthe kinetics of lactose hydrolysis. We observed a significant reduction in the enzymaticactivity of β-GalHis with respect to β-GalWT.Our studies also focus in studying the activity of both β-Gals in the presence ofmultilamellar vesicles (MLVs) of different composition. We conclude that the additionalpositive charges β-GalHis (belonging from histidine residues) promotes the interaction ofthe protein with negatively charged interfaces favoring the effect shown against neutralinterfaces.