Standardization of α-L-iduronidase enzyme assay with Michaelis-Menten kinetics

Li Ou, Tyler L. Herzog, Carrie M. Wilmot, Chester B. Whitley

Research output: Contribution to journalArticlepeer-review

8 Scopus citations

Abstract

The lack of methodological uniformity in enzyme assays has been a long-standing difficulty, a problem for bench researchers, for the interpretation of clinical diagnostic tests, and an issue for investigational drug review. Illustrative of the problem, α-L-iduronidase enzyme catalytic activity is frequently measured with the substrate 4-methylumbelliferyl-α-L-iduronide (4MU-iduronide); however, final substrate concentrations used in different assays vary greatly, ranging from 25μM to 1425μM (Km≈180μM) making it difficult to compare results between laboratories. In this study, α-L-iduronidase was assayed with 15 different substrate concentrations. The resulting activity levels from the same specimens varied greatly with different substrate concentrations but, as a group, obeyed the expectations of Michaelis-Menten kinetics. Therefore, for the sake of improved comparability, it is proposed that α-L-iduronidase enzyme assays should be conducted either (1) under substrate saturating conditions; or (2) when concentrations are significantly below substrate saturation, with results standardized by arithmetic adjustment that considers Michaelis-Menten kinetics. The approach can be generalized to many other enzyme assays.

Original languageEnglish (US)
Pages (from-to)113-115
Number of pages3
JournalMolecular Genetics and Metabolism
Volume111
Issue number2
DOIs
StatePublished - 2014

Bibliographical note

Funding Information:
The authors thank Brenda Koniar for mouse breeding, Renee Cooksley for molecular genetic analysis, Peng Liu for the illustration of Michaelis–Menten kinetics ( Fig. 1 ), Evelyn Redtree for editorial review, and Brenda Diethelm-Okita for regulatory and administrative support. This work was supported by NIH grant P01HD032652 .

Copyright:
Copyright 2014 Elsevier B.V., All rights reserved.

Keywords

  • Hurler syndrome
  • Lineweaver-Burk plot
  • Michaelis-Menten kinetics
  • Mucopolysaccharidosis
  • α-L-iduronidase
  • α-L-iduronidase enzyme assay

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