title: [Role of fibrinogen Bbeta-chain D-region 454-458 residues for assembly and secretion of intact fibrinogen].
abstract: BACKGROUND
  To examine the role of fibrinogen Bbeta-chain D region in the assembly and/or secretion of multichain protein, we synthesized eight variant fibrinogens with truncated Bbeta-chains in the C terminal region, terminating with 454, 455, 456 or 458 residues, and with substitution at Bbeta-455Arg by Lys, Ile, Ala or Asp in Chinese hamster ovary (CHO) cells.
  
  
  METHODS
  A fibrinogen Bbeta-chain expression vector was altered and transfected into CHO cells that expressed normal human fibrinogen Aalpha- and gamma-chains. Expressed fibrinogens of cell lysates and culture media of the established cell lines were subjected to ELISA and immunoblot analysis.
  
  
  RESULTS
  The CHO cells synthesized eight variant Bbeta-chains and assembled these into fibrinogen except for Bbeta-454 and Bbeta-455Asp. However, in the cell lysates, concentrations of these variant fibrinogens were lower than that in wild type cells. These assembled variant fibrinogens were secreted into the culture medium, and the levels in culture media were also lower than that in the medium of wild type cells. Significant differences in the mean ratios of fibrinogen concentration in medium to that in cell lysate were not observed between the variant type cells and the wild type cells.
  
  
  CONCLUSIONS
  Residues of the Bbeta-chain D domain are essential for fibrinogen assembly, especially the Bbeta-455 residue was critical. The present study indicated that the structure of the fibrinogen Bbeta-chain C terminal D region is necessary for fibrinogen assembly, but not for secretion.
authors.affiliation: Department of Biomedical Laboratory Sciences, School of Health Sciences, Shinshu University, Matsumoto 390-8621, Japan. fterasa@shinshu-u.ac.jp
authors.forename: Fumiko
authors.initials: F
authors.lastname: Terasawa
authors.forename: Yuka
authors.initials: Y
authors.lastname: Takezawa
authors.forename: Masako
authors.initials: M
authors.lastname: Hirota-Kawadobora
authors.forename: Nobuo
authors.initials: N
authors.lastname: Okumura
mesh_terms: D000818:Animals; D016466:CHO Cells / Q000378:metabolism; D006224:Cricetinae; D003412:Cricetulus; D003470:Culture Media / Q000737:chemistry; D004797:Enzyme-Linked Immunosorbent Assay; D005340:Fibrinogen / Q000032:analysis / Q000737:chemistry / Q000235:genetics / Q000378:metabolism* / Q000502:physiology; D015151:Immunoblotting; D009154:Mutation; D055503:Protein Multimerization; D017434:Protein Structure, Tertiary
chemical_list: C115499:BBeta fibrinogen; D003470:Culture Media; D005340:Fibrinogen