The Experimentally Determined Velocity of Catalysis could be Higher in the Absence of Sequestration

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Ikechukwu I. Udema
Abraham Olalere Onigbinde


Background: It is not unusual to observe calculated “total” free enzyme ([E]) in enzyme catalysed reaction, but this should include total enzyme-substrate complex ([EST]) which accounts for sequestration.

Objectives: 1) To show indirectly that the velocities of catalytic action can be higher than experimentally observed velocities without sequestration and 2) redefine the relationship between velocity of hydrolysis with Michaelian enzyme and [E], where concentration of substrate, [ST] <  Michaelis-Menten constant, KM.

Methods: A theoretical research and experimentation using Bernfeld method to determine velocities of amylolysis with which to mathematically calculate [EST] and the enzyme-substrate complex ([ES]) prepared for product, P, formation.

Results: The [EST] is < [E]; [EST] and pseudo-first order constant, k decreased with increasing [ST] and increased with increasing concentration of enzyme [ET] while velocity amylolysis, v and maximum velocity of amylolysis, vmax expectedly increased with increasing [ET] and [ST].

Conclusion: The fact is that the [EST] is lower than what is usually referred to as free enzyme ([ET] - [ES]). Therefore, if the additional part of [EST] dissociated into product within the duration of assay, the velocity of amylolysis could be higher. The most important outcome and corollary when [KM] > [ST] is that v a 1/[E], v a [E][ST] and a quadratic relationship exists between pseudo-first order rate constant and maximum velocity of amylolysis; separately, v is not a [E] and if v a [ST] (if 116.PNGv/117.PNG[ST] is constant with coefficient of determination = 1), then KM is not applicable.

Total enzyme-substrate complex, enzyme-substrate complex prepared for product formation, direct proportionality, rate constants, free enzyme, Aspergillus oryzea alpha-amylase.

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How to Cite
Udema, I. I., & Olalere Onigbinde, A. (2019). The Experimentally Determined Velocity of Catalysis could be Higher in the Absence of Sequestration. Asian Journal of Research in Biochemistry, 5(4), 1-12.
Original Research Article


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