1995 Fiscal Year Final Research Report Summary
Function of the formation of SHAP (serum-derived hyaluronan associated protein) -HA complex
| Project/Area Number |
06680594
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| Research Category |
Grant-in-Aid for General Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Research Field |
Structural biochemistry
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| Research Institution | AICHI MEDICAL UNIVERSITY |
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Principal Investigator |
YONEDA Masahiko INSTITUTE FOR NOLECULAR SCIENCE OF MEDICINE,AICHI MEDICAL UNIVERSITY,RESEARCH ASSISTANT, 分子医科学研究所, 助手 (80201086)
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| Project Period (FY) |
1994 – 1995
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| Keywords | hyaluronan / hyaluronan binding protein / inter alpha-trypsin inhibitor / covalent bond / serum / bikunin / extracellular matrix |
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| Research Abstract |
We previously showed that hyaluronan (HA) synthesized by cultured fibloblasts firmly bound serum-derived 85kDa proteins (SHAPs, serum-derived hyaluronan associated proteins). SHAPs were identified with the heavy chains of inter alpha-trypsin inhibitor (ITI) (J.Biol.Chem.268,2672526730,1993). ITI consist of three genetically different peptides, a light chain (bikunin) and two heavy chains (HC1 and HC2). ITI doesn't bind to HA in any assay experiments. We appeared that SHAPs bind covalently to HA.We subjected SHAP-HA complex to limited proteolysis and hyaluronidase-digestion to obtain fragments of the linkage regions. The fragments were analyzed with protein sequencer and electrospray ionization mass spectrometry. The C-terminal Asp of each heavy chain was esterified with C6 hydroxyl group of an internal N-acetylglucosamine of HA chain. This report is the first demonstration to give evidence for the covalent binding of proteins to HA.The reaction of formation requires one of the serum factors (enzymes). It is intersting that the formation of the SHAP-HA complex from HA and ITI is accompanied by the release of bikunin.
The highly metastatic subclone of mouse mammary carcinoma (FM3A P15A) with a high activity in HA synthesis has a large distribution of pericellular HA (HA rich-matrix). Additions of purified ITI have no effect on a size of HA rich-matrix of P15A cultures. Both ITI and the partially purified enzyme added to cultures made it enlarge. It means that the formation of the SHAP-HA complex increases a volume of HA richmatrix. We studied a distribution HA and SHAP in tumor tissues in vivo by immunostaining. We had results that the accumulation of both HA and SHAP in tumor and bikunin localize around tumor. The formation of the SHAP-HA complex on the surface of cells may regulate a construction and size of extracellular matrix and a release of bikunin around cells.
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[Publications] Zhao, M., Yoneda, M., Ohashi, Y., Kurono, S., Iwata, H., Ohnuki, Y., Kimata, K.: "Evidence for the covalent binding of SHAP,heavy chains of inter-alpha-trypsin inhibitor, to hyaluronan." J.Biol.Chem.270. 26657-26663 (1995)
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