PMID: 3766958Aug 15, 1986Paper

A solid-phase substrate of heparanase: its application to assay of human melanoma for heparan sulfate degradative activity

Analytical Biochemistry
M NakajimaG L Nicolson

Abstract

We found a tumor metastasis-associated heparan sulfate (HS)-degrading endoglycosidase in melanoma cells that is a unique endo-beta-glucuronidase (heparanase) capable of specifically cleaving HS at intrachain sites (M. Nakajima, T. Irimura, N. DiFerrante, and G. L. Nicolson, 1984, J. Biol. Chem. 259, 2283-2290). To perform rapid and microscale quantitative assays of heparanase we developed a solid-phase HS substrate by crosslinking radiolabeled HS onto agarose gel beads using one covalent linkage. The HS from bovine lung was partially N-desulfated and labeled with [14C]acetic anhydride. Free HS amino groups were completely acetylated, and reducing terminal saccharides were reductively aminated. The HS derivatives with amino groups at their reducing termini were coupled to amino-reactive agarose beads. Incubation of the solid-phase HS substrates with B16 melanoma cell extracts in the presence of D-saccharic acid 1,4-lactone (a potent exo-beta-glucuronidase inhibitor) resulted in the time- and dose-dependent release of [14C]HS fragments. Human melanoma cell lines were tested for HS-degrading endoglycosidase using the newly developed solid-phase HS substrates. The human malignant melanoma cells tested had high levels of HS-degradin...Continue Reading

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Citations

Oct 8, 2003·Pathology Oncology Research : POR·Dario MarchettiMadhuchhanda Roy
Jul 13, 2004·Analytical Biochemistry·Sachio TsuchidaMasaki Yanagishita
Aug 21, 2003·Analytical Biochemistry·Farhad Behzad, Paul E C Brenchley
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May 12, 2004·Neuro-oncology·Yvonne DenkinsDario Marchetti
Aug 25, 2004·Analytical Biochemistry·Caterina Nardella, Christian Steinkühler
Aug 9, 2016·Biochemical and Biophysical Research Communications·Lipeng WangXiaozhi Wang
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Nov 18, 2018·Molecules : a Journal of Synthetic Chemistry and Natural Product Chemistry·Mohit Chhabra, Vito Ferro
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Feb 1, 1988·Journal of Cellular Biochemistry·M NakajimaG L Nicolson
Feb 24, 2006·Journal of Pharmaceutical and Biomedical Analysis·Koji EnomotoHiroshi Takemoto

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