Deglycosylation of chondroitin sulfate proteoglycan by hydrogen fluoride in pyridine |
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| Authors: | C A Olson R Krueger N B Schwartz |
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| Affiliation: | 1. Pediatric Urology and Regenerative Medicine Research Center, Children’s Hospital Medical Center, Tehran University of Medical Sciences, Tehran, Iran;2. Sina Trauma and Surgery Research Center, Tehran University of Medical Sciences, Tehran, Iran;3. Department of Orthopaedic Surgery, Rothman Institute, Thomas Jefferson University, Philadelphia, PA, USA;4. Toxicology and Poisoning Research Centre, Tehran University of Medical Sciences, Tehran, Iran;5. Department of Toxicology and Pharmacology, Faculty of Pharmacy, Tehran University of Medical Sciences, Tehran, Iran;1. Department of Vascular Surgery, Centre Hospitalier Universitaire Vaudois (CHUV), Lausanne, Switzerland;2. Virginia Tech, Department of Chemistry, Macromolecules Innovation Institute and Virginia Tech Center for Drug Discovery, Blacksburg, VA, USA;1. Organic & Bioorganic Chemistry Laboratory, CSIR-Central Leather Research Institute, Adyar, Chennai-600 020, India;2. Department of Chemistry, Holy Cross College (Autonomous), Tiruchirapalli, 620 002, India;3. Department of Chemical Engineering, Indian Institute of Technology Madras, Chennai 600036, India;4. Biochemistry & Biotechnology Laboratory, CSIR-Central Leather Research Institute, Adyar, Chennai-600020, India;5. Academy of Scientific and Innovative Research (AcSIR), CSIR-CLRI Campus, Adyar, Chennai-600 020, India;1. Department of Environment & Energy Engineering, Chonnam National University, 77 Yongbong-ro, Buk-gu, Gwangju, 61186, Republic of Korea;2. Center for Energy Storage System, Chonnam National University, 77 Yongbong-ro, Buk-gu, Gwangju, 61186, Republic of Korea;3. School of Energy and Chemical Engineering, Ulsan National Institute of Science and Technology, 50 UNIST-gil, Eonyang-eup, Ulju-gun, Ulsan, 44919, South Korea;4. Advanced Materials and Devices Lab, Korea Institute of Energy Research, Daejeon, 305-343, South Korea |
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| Abstract: | The original deglycosylation procedure using HF/pyridine has been modified for maximal removal of carbohydrate from chondroitin sulfate proteoglycan, with minimal alteration of the core protein. Gas-liquid chromatography analysis after treatment for various times showed that 95% of xylose and mannose and 70-85% of other sugars were removed within 30 min, indicating that almost all chondroitin sulfate chains and about 80% of N- and O-linked oligosaccharides were removed. In contrast to the loss of carbohydrate, no change in amino acid composition or loss of immunoreactivity occurred. Longer treatment of up to 16 h resulted in little additional removal of carbohydrate, but did cause a significant decrease in solubility and recovery of the deglycosylated product. Optimal removal of xylose residues after about 1 h was also shown by maximal acceptor activity of the product in a xylosyltransferase assay. Rapid removal of the HF reagent by vacuum evacuation and ion-exchange chromatography, coupled with the reduced time of treatment allowed recovery of an intact, homogenous protein core that is amenable to structural and sequence studies. |
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