The Protein Journal

, Volume 30, Issue 2, pp 143–147 | Cite as

Effect of Human Serum Albumin on the Kinetics of N-glutaryl-L-phenylalanine p-nitroanilide Hydrolysis Catalyzed by α-Chymotrypsin

  • Elsa Abuin
  • Eduardo Lissi
  • Manuel Ahumada
  • Cristian Calderón


The effect of human serum albumin (HSA) addition on the rate of hydrolysis of N-glutaryl-L-phenylalanine p-nitroanilide (GPNA) catalyzed by α-chymotrypsin has been measured in phosphate buffer saline at pH = 7.4. The presence of HSA (up to 200 μM) leads to a decrease in the rate of the process. The reaction follows a Michaelis–Menten mechanism under all the conditions employed. To take into account the effect of substrate depletion due to its binding to albumin ultrafiltration experiments were carried out from which the binding of GPNA to HSA was derived. After correction of the kinetic data taking into account the binding of GPNA to HSA, the activity of the enzyme, and the derived Michaelis constant and catalytic rate constant tends to remain almost independent of the presence of albumin, indicating that the depletion of the substrate due to its binding to HSA is the main factor affecting the enzyme activity.


Enzyme kinetics α-Chymotrypsin N-Glutaryl-L-phenylalanine p-nitroanilide Ultrafiltration 



Human serum albumin


N-glutaryl-L-phenylalanine p-nitroanilide




p-nitrophenyl phosphate


Michaelis constant


Catalytic rate constant


Phosphate buffer saline



Thanks are given to Dicyt (USACH), and Fondecyt (Grant No. 1095036) for financial support. C. Calderon is grateful to Conicyt for a fellowship.


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Copyright information

© Springer Science+Business Media, LLC 2011

Authors and Affiliations

  • Elsa Abuin
    • 1
  • Eduardo Lissi
    • 1
  • Manuel Ahumada
    • 1
  • Cristian Calderón
    • 1
  1. 1.Facultad de Química y BiologíaUniversidad de Santiago de ChileSantiagoChile

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