New strategy for selective and sensitive assay of cathepsin B using a dityrosine-based material

Chan Jin Kim, Dong Ik Lee, Dong Zhang, Chang Ha Lee, Ik Sung Ahn

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7 Citations (Scopus)


The increasing number of reports for disease-related proteases has necessitated materials for the fast, sensitive, and specific assessment of protease activities. The purpose of this study was to synthesize and test a dityrosine-based substrate for the selective assay of a specific cysteine cathepsin. DBDY-Gly-INH)2 was synthesized from the conjugation of N,N′-diBoc-dityrosine (DBDY) with two molecules of glycine and isoniazid (INH) for this purpose. The fluorescence of DBDY (λex = 284-320 nm, λem = 400-420 nm) disappeared due to the quenching effect of INH. However, the protease-catalyzed hydrolysis resulted in the release of INH and recovered the fluorescence of DBDY. When reacted with 13 proteases, DBDY-Gly-INH)2 was hydrolyzed by the cysteine proteases only. Meeting the growing need to discriminate cysteine cathepsins (e.g., cathepsins B, L, and S found at high levels in various cancers), DBDY-Gly-INH)2 was tested as a substrate for cathepsins B, L, and S. Only cathepsin B catalyzed the hydrolysis reaction among the three cathepsins. The reaction rate followed the Michaelis-Menten kinetics, and the KM and kcat/KM values were 2.88 μM and 3.87 × 103 M-1 s-1, respectively, which were comparable to those for the materials reported for the selective assay of cathepsin B. Considering the simple preparation of DBDY-(Gly-INH)2, DBDY-(Gly-INH)2 is believed to be valuable for the sensitive and selective assay of cathepsin B activity.

Original languageEnglish
Pages (from-to)166-173
Number of pages8
JournalAnalytical Biochemistry
Issue number2
Publication statusPublished - 2013

All Science Journal Classification (ASJC) codes

  • Biophysics
  • Biochemistry
  • Molecular Biology
  • Cell Biology


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