Regulation of the catalytic activity and oligomeric composition of enzymes in reversed micelles of surfactants in organic solvents

Abstract

The Phenomenon of the regulation of the catalytic activity of enzymes via changing their oligomeric composition in the system of reversed micelles of sodium bis(2‐ethylhexy)sulfosuccinate (AOT) in octane was studied using z‐chymotrypsin (CT) from bovine brain and alkaline phosphatase (AP) from calf intestinal mucosa. The dependences of the enzyme catalytic activity on the AOT hydration degree (W=[H2O][AOT]), the parameter determining the radius (r 4) of the inner cavity of micelles, usually represent the bell‐shaped curves. The maximal catalytic activity is observed at such Wo when r 4 is equal to the size of the enzyme molecule. The position of this maximum strictly correlates with the enzyme oligomeric composition. Thus, in the case of CT this is observed at Wo =12 when r 1 is equal to the radius (rp ) of the CT globule. In the case of artificially produced conjugate containing six cross‐linked CT molecules, this is observed at W o=43 when r 4 is equal to the radius of the sphere surrounding the absolute octahedron composed of six CT globules. The dependence of the catalytic activity of AP on Wo represent a curve with two maxima that are observed when r 4 is equal to r p of either AP monomer (W=17) or AP dimer (W o=25), Ultracentrifugation experiments revealed that variation of W o causes a change in the oligomeric composition of AP its transition from monomeric (W o<20) to demeric form (W o > 20). Hence, the observed maxima correspond to functioning of different oligomeric forms of AP.

Positive cooperativity in substrate binding of human prostatic acid phosphatase entrapped in AOT–isooctane–water reverse micelles

“…The maximum of the catalytic activity was observed when the size of an inner micelle cavity was equal to that of the solubilized enzyme molecule [17,19,22,27,[29][30][31][32][33][36][37][38][39]. The supposition made on the basis of the results of studies of kinetics was confirmed by sedimentation analysis data.…”

“…In the case of calf intestinal mucosa alkaline phosphatase, two activity maxima have been observed when the radii of the AOT-isooctane-water reverse micellar inner cavity were approximately equal to the sizes of the monomeric and dimeric enzyme forms, respectively. The established widespread opinion that alkaline phosphatase possesses catalytic activity only in dimeric form was established from earlier attempts to separate the subunits of alkaline phosphatase using conventional methods (e.g., in urea solution) that resulted in a loss of the enzyme activity [29,30]. On the other hand, in the case of placental alkaline phosphatase, k cat is growing in an exponential way with w 0 , and no k cat maxima were found [40].…”

“…Several oligomeric enzymes have been reported to dissociate into subunits upon incorporation into reverse micelles under conditions of low w 0 or to associate to higher oligomers in large micelles: alkaline phosphatase [29,30], formate dehydrogenase [31], γ -glutamyl-transferase [32], D-glyceraldehyde-3-phosphate dehydrogenase [30,33], glycogen phosphorylase b [34], lactate dehydrogenase [30,35], malic enzyme [36], penicillin acylase [37,38], and uridine phosphorylase [34,39]. In the case of calf intestinal mucosa alkaline phosphatase, two activity maxima have been observed when the radii of the AOT-isooctane-water reverse micellar inner cavity were approximately equal to the sizes of the monomeric and dimeric enzyme forms, respectively.…”

Carbonic anhydrase activity in Arabidopsis thaliana thylakoid membrane and fragments enriched with PSI or PSII

“…The activity maximum exists because the active enzyme conformation is maintained at specific micelle size that is optimal for a given enzyme. In case of several peaks, each one reflects the functioning of a separate catalytically active unit (Kabanov et al 1991). Thus, the presence of two maxima in the thylakoids implies the presence of at least two species bearing CA activity.…”

pH-dependent Substrate Preference of Pig Heart Lipoamide Dehydrogenase Varies with Oligomeric State

“…It is thought that the close contact between surfactant shell and enzyme helps maintain the latter in the catalytically active conformation. In the case of oligomeric enzymes, the dependence of kinetic parameters on w o shows peaks corresponding to each oligomeric form of an enzyme (49,(51)(52)(53)(54).…”