TY - JOUR
T1 - Development of a high-throughput screening system for the compounds that inhibit collagen-protein interactions
AU - Okano-Kosugi, Hitomi
AU - Matsushita, Osamu
AU - Asada, Shinichi
AU - Herr, Andrew B.
AU - Kitagawa, Kouki
AU - Koide, Takaki
N1 - Funding Information:
This work was supported by Grants-in-Aid for Scientific Research (B) (19390032) and (C) (20590452) from the Japan Society for the Promotion of Science, a Waseda University Grant for Special Research Projects (2008B-098), and the Kagawa University Project Research Fund (2005–2006). We thank T. Matsushita (Nagoya University) for providing the expression vector for the VWF–A3 domain. We also thank T. Nabekura (Niigata University of Pharmacy and Applied Life Sciences) for help with the multiwell plate assay. Appendix A
PY - 2009/11/1
Y1 - 2009/11/1
N2 - Collagen-binding proteins (CBPs) play important roles in various physiological events. Some CBPs are regarded as targets for drug development; for example, platelet glycoprotein VI (GPVI) and heat shock protein 47 (HSP47) are promising targets for the development of novel antiplatelet and antifibrotic drugs, respectively. However, no systematic screening method to search compounds that inhibit collagen-CBP interactions have been developed, and only a few CBP inhibitors have been reported to date. In this study, a facile turbidimetric multiwell plate assay was developed to evaluate inhibitors of CBPs. The assay is based on the finding that CBPs retard spontaneous collagen fibril formation in vitro and that fibril formation is restored in the presence of compounds that interfere with the collagen-CBP interactions. Using the same platform, the assay was performed in various combinations of fibril-forming collagen types and CBPs. This homogeneous assay is simple, convenient, and suitable as an automated high-throughput screening system.
AB - Collagen-binding proteins (CBPs) play important roles in various physiological events. Some CBPs are regarded as targets for drug development; for example, platelet glycoprotein VI (GPVI) and heat shock protein 47 (HSP47) are promising targets for the development of novel antiplatelet and antifibrotic drugs, respectively. However, no systematic screening method to search compounds that inhibit collagen-CBP interactions have been developed, and only a few CBP inhibitors have been reported to date. In this study, a facile turbidimetric multiwell plate assay was developed to evaluate inhibitors of CBPs. The assay is based on the finding that CBPs retard spontaneous collagen fibril formation in vitro and that fibril formation is restored in the presence of compounds that interfere with the collagen-CBP interactions. Using the same platform, the assay was performed in various combinations of fibril-forming collagen types and CBPs. This homogeneous assay is simple, convenient, and suitable as an automated high-throughput screening system.
KW - Collagen
KW - Inhibitor
KW - Peptide
KW - Protein
KW - Screening
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U2 - 10.1016/j.ab.2009.07.017
DO - 10.1016/j.ab.2009.07.017
M3 - Article
C2 - 19615329
AN - SCOPUS:69249209645
SN - 0003-2697
VL - 394
SP - 125
EP - 131
JO - Analytical Biochemistry
JF - Analytical Biochemistry
IS - 1
ER -