ベッコウタケのエンド型とエキソ型ポリガラクツロナーゼの精製とそれらの性質 Purification and Characterization of Endo- and Exo-polygalacturonases from Fomitopsis cytisina

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Author(s)

    • 宮入 一夫 MIYAIRI Kazuo
    • 弘前大学農学生命科学部応用生命工学 Department of Biochemistry and Biotechnology, Faculty of Agriculture and Life Science, Hirosaki University
    • 豊田 素行 TOYODA Motoyuki
    • 弘前大学農学生命科学部応用生命工学 Department of Biochemistry and Biotechnology, Faculty of Agriculture and Life Science, Hirosaki University
    • 奥野 智旦 OKUNO Toshikatsu
    • 弘前大学農学生命科学部応用生命工学 Department of Biochemistry and Biotechnology, Faculty of Agriculture and Life Science, Hirosaki University

Abstract

担子菌ベッコウタケ(Fomitopsis cytisina)の培養濾液から3種類のポリガラクツロナーゼをSDS.PAGE的に均一にまで精製した.これらは反応生成物の分析から,2種類がエンド型(EndoPG I,II),1種類がエキソ型(ExoPG)と決定された.EndoPG I,IIの分子量はSDS-PAGEでともに38kDa,ExoPGは50-60kDaと推定された.これらはいずれも酵素的脱糖鎖によりそれぞれの分子量は37kDaと44kDaに減少した.また,EndoPG I,IIおよびExoPGの等電点はそれぞれ4.3,4.2,4.0であった.いずれの酵素もアミノ酸組成はよく似ていた.また,EndoPG I,IIのN末端20アミノ酸配列には相同性がみられた.これらの至適pHはそれぞれ55,5.0,5.2であり,EndoPG I,IIのpH安定性は4.0から6.0,熱安定性は45℃までであり,ExoPGのpH安定性は3.5から65,熱安定性は45℃までであった.EndoPGIはヘプタ以下のオリゴガラクツロン酸には作用しづらくトリガラクツロン酸にはほとんど作用できなかった.一方,EndoPG IIはIよりオリゴガラクツロン酸によく作用した.本菌のEndoPGはこれまで報告されてきた真菌のものと似ていたが,同じAphyllophorales族のStereum purpureum endoPG IとはKm値,Vmax値,熱安定性が,かなり異なっていた.ExoPGは9GalUA付近の基質に対し最大活性を示し,平均重合度14から42までの基質には最大時の88%程度の活性で一定であった.また,ExoPGは植物起源で報告されていたタイプと同様,不飽和オリゴガラクツロン酸にはまったく作用できなかった.

Three polygalacturonases (PGs) were purified homogeneously in SDS-PAGE from the culture filtrate of white rot basidiomycete, Fomitopsis cytisina . By analysis of reaction products, two of three PGs were concluded to be endo-type (endoPG I and II) and the other to be exo-type (exoPG) . Molecular masses of endoPG I and II were both estimated to be 38 kDa and that of exoPG 50-60 kDa. Their molecular masses by deglycosylating with EndoH decreased by 37, 37 and 44 kDa, respectively. Isoelectric points of endoPG I, II and exoPG were 4.3, 4.2 and 4.0, respectively . Their amino acid compositions were similar to one another and furthermore, endoPG I and II were very similar in the 20 amino acids sequence of N-terminal. Their properties, including their kinetic parameters, were determined. Both endoPG I and II could act more on oligo- and polyGalUAs than hepta of DP and endoPG II could act on tri- and diGalUA. The characteristics of endoPGs from this fungus showed considerable difference from that of Stereum purpureum in spite of allogeneic white rot basidiomycete, being classified under the same Aphyllophorales as S. purpureum. ExoPG had the highest activity on nonaGalUA and, like plants exoPG, could not act on 4, 5-unsatd. oligoGalUA.

Journal

  • Journal of Applied Glycoscience

    Journal of Applied Glycoscience 48(2), 105-114, 2001-04-01

    The Japanese Society of Applied Glycoscience

References:  24

Cited by:  1

Codes

  • NII Article ID (NAID)
    10008252473
  • NII NACSIS-CAT ID (NCID)
    AN10453916
  • Text Lang
    ENG
  • Article Type
    Journal Article
  • ISSN
    13403494
  • NDL Article ID
    5764208
  • NDL Source Classification
    ZP24(科学技術--化学・化学工業--糖・澱粉)
  • NDL Call No.
    Z17-15
  • Data Source
    CJP  CJPref  NDL  J-STAGE  JASI 
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