Cloning and bacterial expression systems for recombinant human heparanase production: Substrate specificity investigation by docking of a putative heparanase substrate. (2nd October 2017)

• Record Type: Journal Article
• Authors: Pennacchio, Angela; Capo, Alessandro; Caira, Simonetta; Tramice, Annabella; Varriale, Antonio; Staiano, Maria; D'Auria, Sabato
• Other Names: Dainese Enrico guestEditor.; Gilardi Gianfranco guestEditor.; Maccarrone Mauro guestEditor.
• Is Part Of: Biotechnology and applied biochemistry. Volume 65:Number 1(2018)
• Page Start: 89
• Page End: 98
• Publication Date: 2017-10-02
• Subjects: activity -- docking -- fluorogenic sulfate substrate -- human heparanase -- interaction ligand -- quantitative assay; Biotechnology -- Periodicals; Biochemical engineering -- Periodicals; Biochemistry -- Periodicals; Biochemistry -- Periodicals; Genetic Techniques -- Periodicals; Microbiological Techniques -- Periodicals; 660.6
• DOI: 10.1002/bab.1582
• Languages: English
• ISSNs: 0885-4513

Abstract

Abstract: Human heparanase (HPSE) is an enzyme that degrades the extracellular matrix. It is implicated in a multiplicity of physiological and pathological processes encouraging angiogenesis and tumor metastasis. The protein is a heterodimer composed of a subunit of 8 kDa and another of 50 kDa. The two protein subunits are noncovalently associated. The cloning and expression of the two protein subunits in Escherichia coli and their subsequent purification to homogeneity under native conditions result in the production of an active HPSE enzyme. The substrate specificity of the HPSE was studied by docking of a putative substrate that is a designed oligosaccharide with the minimum recognition backbone, with the additional 2‐N‐sulfate and 6‐O‐sulfate groups at the nonreducing GlcN and a fluorogenic tag at the reducing extremity GlcN. To develop a quantitative fluorescence assay with this substrate would be extremely useful in studies on HPSE, as the HPSE cleavage of fluorogenic tag would result in a measurable response.