title: Kinetics and thermodynamic transitions of N-acetyl-beta-D-glucosaminidase A and B in free and bound forms: role of cellulose ion-exchangers.
abstract: The kinetic and thermodynamic properties of N-acetyl-beta-D-glucosaminidase A (Hex A) and N-acetyl-beta-D-glucosaminidase beta (Hex B) from goat testes were investigated in free and bound (after binding them on ion-exchangers such as DEAE- or CM-cellulose respectively) forms. The optimum pH of free Hex A and Hex B was at 4.2 and 5.4, whereas the bound forms showed the optimum pH at 4.0 and 5.2 respectively. While apparent Km of free and bound Hex A (0.8 and 1.0 mM respectively) did not differ, the Km of Hex B increased when bound on CM-cellulose (Km of free Hex B = 0.96 mM versus bound Hex B = 1.6 mM). Though the free Hex A was more thermo-labile than the free Hex B, both isozymes, on insoluble matrices decayed at faster rates on heating. Activation analysis revealed that the energy of activation (Eoa) for transition state of free Hex B (81 Kcal deg-1 mole-1) did not differ from Eoa of bound Hex B. On the other hand, Eoa of free Hex A declined from 77.2 to 71.1 Kcal deg-1 mole-1 when heat transitions were carried out in free and bound state respectively. Thermodynamic analysis suggested a change in entropy of activation (delta S) of free Hex A and Hex B as 200 and 211 eu respectively. While delta S of Hex B did not change after heat transitions, delta S of Hex A was 182.5 eu.
authors.affiliation: Department of Biophysics, Panjab University, Chandigarh, India.
authors.forename: G S
authors.initials: GS
authors.lastname: Gupta
authors.forename: R
authors.initials: R
authors.lastname: Raina
mesh_terms: D000818:Animals; D003636:DEAE-Cellulose; D006041:Goats; D054818:Hexosaminidase A; D054819:Hexosaminidase B; D006863:Hydrogen-Ion Concentration; D007474:Ion Exchange; D007527:Isoenzymes; D007700:Kinetics; D008297:Male; D013737:Testis / Q000201:enzymology; D013816:Thermodynamics; D001619:beta-N-Acetylhexosaminidases / Q000302:isolation & purification / Q000378:metabolism*
chemical_list: D007527:Isoenzymes; D003636:DEAE-Cellulose; D054818:Hexosaminidase A; D054819:Hexosaminidase B; D001619:beta-N-Acetylhexosaminidases