Interfasial behavior of β -galactosidases

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

Rosario

Encuentro; L Reunión Anual de la Sociedad Argentina de Biofísica; 2022

Sociedad Argentina de Biofísica

β-galactosidase (β-Gal) is an enzyme that hydrolyzes lactose, its natural substrate. This lactase is ofgreat biotechnological interest and is used in the dairy industry, pharmacology and molecularbiology. In our laboratory we are interested in finding the optimal environmental condition toimprove the structure-function of the protein.Currently, we produce a recombinant β-Galactosidase (β-Gal His ) with 6 histidine residues in thecarboxyl terminus that facilitates its subsequent purification by metal ion affinity chromatography.In this work we tested the protein in heterogeneous systems such as multilamellar vesicles (MLVs)dispersions and monomolecular layers of different lipid composition (pure Egg-PC, DOPG and in amixture of Egg-PC-DOPG (ratio 80:20)).We compare the catalytic activity against lactose and the thermal stability of β-Gal His and its nativecounterpart (β-Gal WT ) in the presence and absence of multilamellar vesicles (MLVs) dispersion. Inaddition, we evaluated the ability of the enzyme to penetrate to the lipid monomolecular layers.β-Gal His present lower activity and thermal stability than β-Gal WT that could be explained by aparticular conformational evidenced by the intrinsic fluorescence of the protein. Our resultsdemonstrated that both enzymes, but mainly the β-Gal His , tended to be activated andthermoprotected up to 50ºC by MLVs. These results are in accordance with the determination of theTm values. The fact that β-Gal His was highly activated by neutral PC (Egg-PC) could be related withthe highest penetration of β-Gal His to Egg-PC monomolecular layers.