title: Enzymatic synthesis of hyaluronan hybrid urinary trypsin inhibitor.
abstract: Human urinary trypsin inhibitor is a proteoglycan that has a single low-sulfated chondroitin 4-sulfate chain at the seryl residue in position 10 of the core protein as a glycosaminoglycan moiety, and is used as an anti-inflammatory medicine based on the protease inhibitory activity of the core protein. However, the functions of the glycosaminoglycan moiety have not yet been elucidated in detail. In the present study, the glycosaminoglycan chains of a native urinary trypsin inhibitor were remodeled to hyaluronan chains, with no changes to the core protein, using transglycosylation as a reverse reaction of the hydrolysis of bovine testicular hyaluronidase, and the properties of the hybrid urinary trypsin inhibitor were then analyzed. The trypsin inhibitory activitiy of the hyaluronan hybrid urinary trypsin inhibitor was similar to that of the native type; however, its inhibitory effect on the hydrolysis of hyaluronidase were not as strong as that of the native type. This result demonstrated that the native urinary trypsin inhibitor possessed hyaluronidase inhibitory activity on its chondroitin sulfate chain. The hyaluronan hybrid urinary trypsin inhibitors obtained affinity to a hyaluronan-binding protein not exhibited by the native type. The interactions between the hyaluronan hybrid urinary trypsin inhibitors and phosphatidylcholine (abundant in the outer layer of plasma membrane) were stronger than that of the native type. Hyaluronan hybrid urinary trypsin inhibitors may be useful for investigating the functions of the glycosaminoglycan chains of urinary trypsin inhibitors and hyaluronan, and our hybrid synthesizing method may be used widely in research for future medical applications.
authors.affiliation: Department of Glycotechnology, Center for Advanced Medical Research, Hirosaki University Graduate School of Medicine, 5 Zaifu-cho, Hirosaki 036-8562, Japan; Department of Biochemistry, Hirosaki University School of Medicine, 5 Zaifu-cho, Hirosaki 036-8562, Japan. Electronic address: kaki@hirosaki-u.ac.jp.
authors.forename: Ikuko
authors.initials: I
authors.lastname: Kakizaki
authors.affiliation: Wakamoto Pharmaceutical Co. Ltd., 378 Kanate Ooimachi Ashigarakamigun, Kanagawa 258-0018, Japan.
authors.forename: Ryoki
authors.initials: R
authors.lastname: Takahashi
authors.affiliation: Department of Physics, Graduate School of Sciences, Kyushu University, 6-10-1 Hakozaki, Higashi-ku, Fukuoka 812-8581, Japan.
authors.forename: Miho
authors.initials: M
authors.lastname: Yanagisawa
authors.affiliation: Department of Physics, Graduate School of Sciences, Kyushu University, 6-10-1 Hakozaki, Higashi-ku, Fukuoka 812-8581, Japan.
authors.forename: Futaba
authors.initials: F
authors.lastname: Yoshida
authors.affiliation: Department of Biochemistry, Hirosaki University School of Medicine, 5 Zaifu-cho, Hirosaki 036-8562, Japan.
authors.forename: Keiichi
authors.initials: K
authors.lastname: Takagaki
mesh_terms: D000268:Adhesiveness; D000818:Animals; D002417:Cattle; D002809:Chondroitin Sulfates / Q000737:chemistry; D006023:Glycoproteins / Q000138:chemical synthesis* / Q000737:chemistry* / Q000494:pharmacology; D006820:Hyaluronic Acid / Q000737:chemistry*; D006821:Hyaluronoglucosaminidase / Q000378:metabolism*; D006868:Hydrolysis; D020128:Inhibitory Concentration 50; D014361:Trypsin Inhibitors / Q000138:chemical synthesis* / Q000737:chemistry* / Q000494:pharmacology
chemical_list: D006023:Glycoproteins; D014361:Trypsin Inhibitors; D006820:Hyaluronic Acid; D002809:Chondroitin Sulfates; D006821:Hyaluronoglucosaminidase; C028665:urinastatin
keywords: Chondroitin sulfate; Glycosaminoglycan; Hyaluronan; Hyaluronidase; Transglycosylation; Urinary trypsin inhibitor