Studies of thiamin diphosphate binding to the yeast apotransketolase

• Published in: Journal of molecular catalysis. B, Enzymatic Vol. 26; no. 1; pp. 33 - 40
• Main Authors: Selivanov, Vitaliy A, Kovina, Marina V, Kochevova, Natalia V, Meshalkina, Ludmilla E, Kochetov, German A
• Format: Journal Article
• Language: English
• Published: Elsevier B.V 03.11.2003
• Subjects: Kinetic modeling; Thiamin diphosphate; Transketolase; Thiamin diphosphate; Kinetic modeling; Transketolase; ThDP
• DOI: 10.1016/S1381-1177(03)00115-2
• ISSN: 1381-1177

Previously it was shown that the binding of thiamin diphosphate proceeds through two steps: fast primary binding and the subsequent slow conformational transition of the apoprotein. In the presence of Ca 2+, the coenzyme binding occurs with negative cooperativity—owing to the increased rate of the reverse conformational transfer in one of the active centers after completion of ThDP binding at both active centers. There are three viewpoints on the enzyme behavior upon replacement of Ca 2+ with Mg 2+: (a) negative cooperativity between the two centers is retained; (b) turns positive; (c) totally disappears. In this work, a comparative investigation of the interaction between ThDP and apotransketolase was undertaken and the negative cooperativity between the two centers in the presence of Mg 2+, just as in the presence of Ca 2+ was demonstrated—albeit with the former cation it was somewhat less pronounced. The negative cooperativity with Mg 2+, just as with Ca 2+, was caused by an increase in the rate of reverse conformational transfer after the ThDP binding completion in both active centers.