title: Relationship between length variations in Ser/Asp-rich repeats in phosphophoryn and in vitro precipitation of calcium phosphate.
abstract: OBJECTIVE
  Phosphophoryn (PP) is generated from the proteolytic cleavage of dentin sialophosphoprotein (DSPP). PP which contains tandem serine/asparatic acid rich repeats (SDrr) is known to enhance dentin mineralization. The nucleotide sequences coding SDrr are identified in the DSPP genes of toothed animals and the length variations of SDrr between intra- and inter-species have been reported. However, it remains unknown about the relationship between the length variations in SDrr and the functions of PP in matrix mineralization.
  
  
  METHODS
  By utilizing a mammalian expression system, we generated several recombinant PP proteins (rPP) containing SDrr of different lengths and analyzed their effects on the precipitation of calcium phosphate with an in vitro gel diffusion system.
  
  
  RESULTS
  rPP-Δ37.6 SDrr and rPP-Δ63.5 SDrr, which possessed shortened SDrr that accounted for 62.4 and 36.5% the length of SDrr in full-length rPP (rPP-full), respectively, induced the precipitation of calcium phosphate similar to that of rPP-full at the same molar concentration, whereas rPP-ΔSDrr, in which SDrr were flipped, did not. Furthermore, rPP-Δ63.5 SDrr significantly increased the accumulation of calcium compared with rPP-full at adjusted concentrations so that the same amounts of SDrr were embedded. The results of an ELISA analysis indicated that the amounts of rPP-Δ37.6 SDrr and rPP-Δ63.5 SDrr secreted from transfected cells were 5.2- and 7.1-fold greater than that of rPP-full, respectively.
  
  
  CONCLUSIONS
  The generated rPP-Δ63.5 SDrr which can be substituted for rPP-full may be a candidate for a therapeutic molecule to facilitate hard tissue generation such as reparative dentin formation.
authors.affiliation: Department of Biological Endodontics, Integrated Health Sciences, Institute of Biomedical and Health Sciences, Hiroshima University, Hiroshima, 734-8553, Japan.
authors.forename: Seiji
authors.initials: S
authors.lastname: Kobuke
authors.affiliation: Department of Biological Endodontics, Integrated Health Sciences, Institute of Biomedical and Health Sciences, Hiroshima University, Hiroshima, 734-8553, Japan. Electronic address: suzukis@hiroshima-u.ac.jp.
authors.forename: Shigeki
authors.initials: S
authors.lastname: Suzuki
authors.affiliation: Department of Biological Endodontics, Integrated Health Sciences, Institute of Biomedical and Health Sciences, Hiroshima University, Hiroshima, 734-8553, Japan.
authors.forename: Hiroaki
authors.initials: H
authors.lastname: Hoshino
authors.affiliation: Section of Orthodontics and Dentofacial Orthopedics, Faculty of Dental Science, Kyushu University, Fukuoka, 812-5852, Japan.
authors.forename: Naoto
authors.initials: N
authors.lastname: Haruyama
authors.affiliation: Section of Periodontology, Division of Oral Rehabilitation, Faculty of Dental Science, Kyushu University, Fukuoka, 812-5852, Japan.
authors.forename: Fusanori
authors.initials: F
authors.lastname: Nishimura
authors.affiliation: Department of Biological Endodontics, Integrated Health Sciences, Institute of Biomedical and Health Sciences, Hiroshima University, Hiroshima, 734-8553, Japan.
authors.forename: Hideki
authors.initials: H
authors.lastname: Shiba
mesh_terms: D000818:Animals; D001224:Aspartic Acid / Q000737:chemistry*; D001483:Base Sequence; D015153:Blotting, Western; D002130:Calcium Phosphates / Q000737:chemistry*; D011232:Chemical Precipitation; D003804:Dentin / Q000737:chemistry*; D004797:Enzyme-Linked Immunosorbent Assay; D016326:Extracellular Matrix Proteins; D066298:In Vitro Techniques; D008322:Mammals / Q000235:genetics; D010750:Phosphoproteins / Q000737:chemistry* / Q000235:genetics*; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry; D012694:Serine / Q000737:chemistry*; D012795:Sialoglycoproteins; D020080:Tandem Repeat Sequences; D014074:Tooth Calcification / Q000502:physiology*; D014162:Transfection; D000099311:Dentin Sialophosphoprotein
chemical_list: D001224:Aspartic Acid; D002130:Calcium Phosphates; D016326:Extracellular Matrix Proteins; D010750:Phosphoproteins; D011994:Recombinant Proteins; D012694:Serine; D012795:Sialoglycoproteins; D000099311:Dentin Sialophosphoprotein; C017630:phosphophoryn; C020243:calcium phosphate
keywords: In vitro gel diffusion system; Phosphophoryn (PP); Recombinant protein preparation; Serine/aspartic acid-rich repeats (SDrr)