BioChip Journal

, Volume 10, Issue 3, pp 198–207 | Cite as

Development of a matrix metalloproteinase-2 (MMP-2) biosensing system by integrating an enzyme-mediated color development reaction into a common electronics components setup

  • Cun Qiang Zhang
  • Yoo Min Park
  • Dokyung Yang
  • Tae Hyeon YooEmail author
  • Hyun C. YoonEmail author
Original Article


Matrix metalloproteinase-2 (MMP-2) is closely related to the proliferation and invasion of various types of cancers. The protease is secreted by malignant tumor cells, thus allowing the enzyme to serve as a biomarker for cancer diagnosis. Methods have been developed to analyze MMP-2 activities; however, their applications to disease diagnosis have not been widely demonstrated yet because of the need for highend analytical equipment and labor-intensive processes. In this study, we developed an MMP-2 activity assay system by integrating an engineered autoinhibited β-lactamase which can be activated by MMP-2 in an optical sensing system consisting of reassembled common electronic components, such as a laser diode, a solar cell, and a multimeter. The autoinhibited β-lactamase was immobilized on a polymeric biosensing channel by a polydopamine coating and self-assembled monolayer methods. In the presence of MMP-2, the immobilized autoinhibited β-lactamase was converted to an active form that hydrolyzed the chromogenic cephalosporin CENTA, thereby changing the substrate color from pale yellow (λmax=340 nm) to highly discernible chrome yellow (λmax=405 nm). By reading the interfered laser-light intensity, we were able to analyze MMP-2 activities precisely both with the samples prepared in a buffer solution and also those in urine. These results suggested that the developed system can be used for the quantitative analysis of enzyme activity related to cancer diagnosis.


Matrix metalloproteinase-2 Autoinhibited β-lactamase Optical biosensor 


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  1. 1.
    Aaronson, S.A. Growth factors and cancer. Science 254, 1146–1153 (1991).CrossRefGoogle Scholar
  2. 2.
    Duffy, M.J. The role of proteolytic enzymes in cancer invasion and metastasis. Clin. Exp. Metastasis. 10, 145–155 (1992).CrossRefGoogle Scholar
  3. 3.
    Rosenberg, S.A. A New Era for Cancer Immunotherapy Based on the Genes that Encode Cancer Antigens. Immunity. 10, 281–287 (1999).CrossRefGoogle Scholar
  4. 4.
    Westermarck, J. & Kähäri, V.M. Regulation of matrix metalloproteinase expression in tumor invasion. FASEB. J. 13, 781–792 (1999).Google Scholar
  5. 5.
    Roy, R., Yang, J. & Moses, M.A. Matrix metalloproteinases as novel biomarkers and potential therapeutic targets in human cancer. J. Clin. Oncol. 27, 5287–5297 (2009).CrossRefGoogle Scholar
  6. 6.
    Zucker, S. & Vacirca, J. Role of matrix metalloproteinases (MMPs) in colorectal cancer. Cancer. Metastasis. Rev. 23, 101–117 (2004).CrossRefGoogle Scholar
  7. 7.
    Turpeenniemi-Hujanen, T. Gelatinases (MMP-2 and -9) and their natural inhibitors as prognostic indicators in solid cancers. Biochimie. 87, 287–297 (2005).CrossRefGoogle Scholar
  8. 8.
    Moses, M.A., Wiederschain, D., Loughlin, K.R., Zurakowski, D. & Freeman, M.R. Increased Incidence of Matrix Metalloproteinases in Urine of Cancer Patients. Cancer. Res. 58, 1395–1399 (1998).Google Scholar
  9. 9.
    Angelina, D.C., Daniela, T., Angela, M. & Vincenzo, M. Urinary gelatinase activities (matrix metalloproteinases 2 and 9) in human bladder tumors. Oncol. Rep. 15, 1321–1326 (2006).CrossRefGoogle Scholar
  10. 10.
    Angelina, D.C., Angela, M., Daniela, T. & Matteo, F. Matrix metalloproteinase-2 and -9 in the urine of prostate cancer patients. Oncol. Rep. 24, 3–8 (2010).Google Scholar
  11. 11.
    Coticchia, C.M., Curatolo, A.S., Zurakowski, D., Yang, J. & Daniels, K.E. Urinary MMP-2 and MMP-9 predict the presence of ovarian cancer in women with normal CA125 levels. Gynecol. Oncol. 123, 295–300 (2011).CrossRefGoogle Scholar
  12. 12.
    Fitzsimmons, P.J. et al. Urinary levels of matrix metalloproteinase 9 and 2 and tissue inhibitor of matrix metalloproteinase in patients with coronary artery disease. J. Atheroscler. Res. 194, 196–203 (2007).CrossRefGoogle Scholar
  13. 13.
    Kleber, C.J., Spiess, A., Kleber, J.B., Hinz, U. & Weiss, J. Urinary matrix metalloproteinases-2/9 in healthy infants and haemangioma patients prior to and during propranolol therapy. Eur. J. Pediatr. 71, 941–946 (2012).CrossRefGoogle Scholar
  14. 14.
    Eissa, S. et al. Noninvasive Diagnosis of Bladder Cancer by Detection of Matrix Metalloproteinases (MMP-2 and MMP-9) and Their Inhibitor (TIMP-2) in Urine. Eur. Urol. 52, 1388–1397 (2007).CrossRefGoogle Scholar
  15. 15.
    Wysocki, A.B., Staiano-Coico, L. & Grinnell, F. Wound Fluid from Chronic Leg Ulcers Contains Elevated Levels of Metalloproteinases MMP-2 and MMP-9. J. Invest. Dermatol. 101, 64–68 (1993).CrossRefGoogle Scholar
  16. 16.
    Wang, C. et al. Expression and characterization of common carp (Cyprinus carpio) matrix metalloproteinase-2 and its activity against type I collagen. J. Biotechnol. 177, 45–52 (2014).CrossRefGoogle Scholar
  17. 17.
    McQuibban, G.A., Butler, G.S., Gong, J.H. & Bendall, L. Matrix Metalloproteinase Activity Inactivates the CXC Chemokine Stromal Cell-derived Factor-1. J. Biol. Chem. 276, 43503–43508 (2001).CrossRefGoogle Scholar
  18. 18.
    Yang, J. et al. Detection of MMP activity in living cells by a genetically encoded surface-displayed FRET sensor. BBA-Mol. Cell. Res. 1773, 400–407 (2007).Google Scholar
  19. 19.
    Wang, Y.H., Shen, P., Li, C.Y., Wang, Y.Y. & Liu, Z.H. Upconversion Fluorescence Resonance Energy Transfer Based Biosensor for Ultrasensitive Detection of Matrix Metalloproteinase-2 in Blood. Anal. Chem. 84, 1466–1473 (2012).CrossRefGoogle Scholar
  20. 20.
    Kim, H.J., Yoon, H.K. & Yoo, T.H. Engineering b-lactamase zymogens for use in protease activity assays. Chem. Commun. 50, 10155–10157 (2014).CrossRefGoogle Scholar
  21. 21.
    Han, Y.D., Chun, H.J. & Yoon, H.C. The transformation of common office supplies into a low-cost optical biosensing platform. Biosens. Bioelectron. 59, 259–268 (2014).CrossRefGoogle Scholar
  22. 22.
    Carine, B., Catherine, M., Florence, M. & Sandrine, R. CENTA as a Chromogenic Substrate for Studying ß-Lactamases. Antimicrob. Agents Chemother. 45, 1868–1871 (2001).CrossRefGoogle Scholar
  23. 23.
    Smith, E.R., Zurakowski, D., Saad, A. & Scott, R.M. Urinary Biomarkers Predict Brain Tumor Presence and Response to Therapy. Clin. Cancer. Res. 14, 2378–2386 (2008).CrossRefGoogle Scholar
  24. 24.
    Roy, R. et al. Urinary TIMP-1 and MMP-2 levels detect the presence of pancreatic malignancies. Br. J. Cancer. 111, 1772–1779 (2014).CrossRefGoogle Scholar
  25. 25.
    International Conference on Harmonisation of Technical Requirements for Registration of Pharmaceuticals for Human Use, ICH Harmonised Tripartite Guideline, Validation of analytical procedures: Text and Methodology Q2(R1) 2005.Google Scholar
  26. 26.
    Laraki, N., Franceschini, N., Roßsolini, G.M., Santucci, P. & Meunier, C. Biochemical Characterization of the Pseudomonas aeruginosa 101/1477 Metallo-Lactamase IMP-1 Produced by Escherichia coli. Antimicrob. Agents Chemother. 43, 902–906 (1999).Google Scholar

Copyright information

© The Korean BioChip Society and Springer-Verlag Berlin Heidelberg 2016

Authors and Affiliations

  1. 1.Department of Molecular Science & TechnologyAjou UniversitySuwonKorea

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