Effect of partitioning equilibria on the activity of β-galactosidase in heterogeneous media

Molecular Packing Tunes the Activity of Kluyveromyces lactis β-Galactosidase Incorporated in Langmuir−Blodgett Films

“…Subindexes indicate the initial surface pressure (π i ) in mN/m. The protein subphase concentration was 77.6 μg/mL. b Approximate equilibrium surface pressure of bilayers. c Calculated considering a 1 mg/mL lipid concentration, 1 μm diameter MLVs that exposed 25% of the total lipid, a dpPC molecular area of 70 Å 2 at the equilibrium surface pressure of a bilayers and a membrane−water partition coefficient for β-Gal P m/w = 1122, according to data taken from ref . d They might be aggregates; hence, they should be expressed as spots/Å 2 . e From the Stokes radio of K. lactis β-Gal in the native and the unfolded states (52 and 147 Å, respectively) its area/molecule is 8500 and 6.7 × 10 4 Å 2 /molecules, respectively. On the other hand, smaller ( r = 500 Å) and bigger ( r = 2000 Å) spots exhibited average surface areas of 0.780 × 10 6 and 1.25 × 10 7 Å 2 /spot, respectively; hence, the mean molecular density per spot in LB 35 is ca.…”

“… c Calculated considering a 1 mg/mL lipid concentration, 1 μm diameter MLVs that exposed 25% of the total lipid, a dpPC molecular area of 70 Å 2 at the equilibrium surface pressure of a bilayers and a membrane−water partition coefficient for β-Gal P m/w = 1122, according to data taken from ref . …”

“…In these kinds of systems the kinetics of enzyme-catalyzed reactions may be modulated through several mechanisms affecting, e.g., the distribution of enzymes and/or substrates between the different compartments and the supramolecular organization of the substrate (in the case of amphipathic and self-organized substrates). − The accompanying molecular crowding may be affecting the probability of enzyme−substrate collisions due to the excluded volume and the thermodynamic activity of water due to its structuring at surfaces . The enzyme binding to a nonsubstrate surface can modulate the enzyme conformation and activity. − Moreover, the reaction confinement to dimensionally restricted spaces such as a percolation matrix, channel, or surface can cause significant effects on the reaction mechanism and rate. ,− …”

“…In previous works we demonstrated that β-galactosidase from Escherichia coli (β-Gal), which is a soluble protein, could be modulated in heterogeneous media. This modulation was dependent on the enzyme interaction with lipid/water interfaces and exerted at different levels, e.g., the enzyme conformation and reaction mechanism included an apparent effect associated with the partition of one of the reaction products ( o -nitrophenol) between the membrane and the water phases . Localization of this substance within the polar headgroup region of the membrane also played a significant role in the modulation of the hydrolytic reaction mechanism.…”

Composition-driven Surface Domain Structuring Mediated by Sphingolipids and Membrane-active Proteins

“…The lipid surface packing, interfacial hydration and molecular crowding affect the activity of b-galactosidase [277]. Also, the activity of The arrows indicate beginning of steadystate regime.…”