2024-03-28T22:07:43Zhttps://eprints.lib.hokudai.ac.jp/dspace-oai/requestoai:eprints.lib.hokudai.ac.jp:2115/442072022-11-17T02:08:08Zhdl_2115_20055hdl_2115_8527Structure of Pleiotrophin- and Hepatocyte Growth Factor-binding Sulfated Hexasaccharide Determined by Biochemical and Computational ApproachesLi, FuchuanNandini, Chilkunda D.Hattori, TomohideBao, XingfengMurayama, DaisukeNakamura, ToshikazuFukushima, NobuhiroSugahara, Kazuyuki460Endogenous pleiotrophin and hepatocyte growth factor (HGF) mediate the neurite outgrowth-promoting activity of chondroitin sulfate (CS)/dermatan sulfate (DS) hybrid chains isolated from embryonic pig brain. CS/DS hybrid chains isolated from shark skin have a different disaccharide composition, but also display these activities. In this study, pleiotrophin- and HGF-binding domains in shark skin CS/DS were investigated. A high-affinity CS/DS fraction was isolated using a pleiotrophin-immobilized column. It showed marked neurite outgrowth-promoting activity and strong inhibitory activity against the binding of pleiotrophin to immobilized CS/DS chains from embryonic pig brain. The inhibitory activity was abolished by chondroitinase ABC or B, and partially reduced by chondroitinase AC-I. A pentasulfated hexasaccharide with a novel structure was isolated from the chondroitinase AC-I digest using pleiotrophin-affinity and anion-exchange chromatographies. It displayed a potent inhibitory effect on the binding of HGF to immobilized shark skin CS/DS chains, suggesting that the pleiotrophin- and HGF-binding domains at least partially overlap in the CS/DS chains involved in the neuritogenic activity. Computational chemistry using molecular modeling and calculations of the electrostatic potential of the hexasaccharide and two pleiotrophin-binding octasaccharides previously isolated from CS/DS hybrid chains of embryonic pig brain identified an electronegative zone potentially involved in the molecular recognition of the oligosaccharides by pleiotrophin. Homology modeling of pleiotrophin based on a related midkine protein structure predicted the binding pocket of pleiotrophin for the oligosaccharides and provided new insights into the molecular mechanism of the interactions between the oligosaccharides and pleiotrophin.Journal Articleapplication/pdfhttp://hdl.handle.net/2115/44207https://eprints.lib.hokudai.ac.jp/dspace/bitstream/2115/44207/1/revisedtextLisubmitted100605MZ.pdf0021-9258Journal of Biological Chemistry2853627673276852010-09-03enginfo:pmid/20584902info:doi/10.1074/jbc.M110.118703author