Microbiology and Biotechnology Letters (한국미생물·생명공학회지)

The Korean Society for Microbiology and Biotechnology (한국미생물·생명공학회)
권호 표지

Chemical Modification of the $\beta$-D-Xylosidase from Bacillus stearothermophilus

화학적 수식에 의한 Bacillus stearothermophilus $\beta$-D-Xylosidase 의 연구

  • 서정한 (고려대학교 자연자원대학 유전공학과) ;
  • 최용진 (고려대학교 자연자원대학 유전공학과)
  • Published : 1994.12.01
Download PDF
⟨ Previous Next ⟩

Abstract

Essential amino acids involving in the catalytic mechanism of the $\beta$-D-xylosidase of Bacillus stearothermophilus were determined by chemical modification studies. Among various che- mical modifiers tested N-bromosuccinimide (NBS), $\rho$-hydroxymercurybenzoate (PHMB), N-ethylma- leimide, 1-[3-(di-ethylamino)-propyl]$-3-ethylcarbodi-imide (EDC), and Woodward's Reagent K(WRK)inactivated the enzyme, resulting in the residual activity of less than 20%. WRK reduced the enzyme activity by modifying carboxylic amino acids, and the inactivation reacion proceeded in the form of pseudo-first-order kinetics. The double-lagarithmic plot of the observed pseudo-first- order rate constant against the modifier concentration yielded a reaction order of 2, indicating that two carboxylic amino acids were essential for the enzyme activity. The $\beta$-D-xylosidase was also inactivated by N-ethylmaleimide which specifically modified a cysteine residue with a reaction order of 1, implying that one cysteine residue was important for the enzyme activity. Xylobiose protected the enzyme against inactivation by WRK and N-ethylmaleimide, revealing that carboxylic amino acids and a cysteine residue were present at the substrate-binding site of the enzyme molecule.

Keywords

References

  1. Adv. Carbohydr. Chem. Biochem. v.32 Hemicellulases, their occurence, purification, properties and applications Dekker,R.F.H.;G.N.Richards
  2. Trends Biotechnol. v.3 Microbial xylanolytic systems Biely,P.
  3. J. Sci. Food Agri. v.34 Substituent groups linked by alkali-labile bonds to arabinose and xylose residues of legume, grass, and cereal straw cell walls and their fate during digestion by rumen microorganisms Chesson,A.;A.H.Gordon;J.A.Lomax
  4. Kor. J. Appl. Microbiol. Tech. v.17 Production of xylanase by Bacillus stearothermophilus Song,H.S.;Y.J.Choi
  5. Kor. J. Appl. Microbiol. Tech. v.20 Molecular cloning and expression of an endo-xylanase gene from Bacillus stearothermophilus into Escherichia coli Cho,S.G.;S.S.park;Y.I.Park;Y.J.Choi
  6. Kor. J. Appl. Microbiol. Tech. v.21 Molecular cloning and expression of the acetyl xylan esterases gene of Bacillus stearothermophilus in Escherichia coli Kim,I.S.;S.G.Cho;Y.J.Choi
  7. Kor. J. Appl. Microbiol. Biotech v.20 Molecular cloning and expression of bacillus stearothermophilus β-D-xylosidase gene in E. coli Oh,S.W.;S.S.Park;Y.J.Choi
  8. Kor. J. Appl. Microbiol Biotech. v.20 Purification and characteriztion of exo-xylanase from Escherichia coli harboring the recombinant plasmid pMG1 Mun,A.R.;Y.J.Choi
  9. Kor. J. Appl. Microbiol Biotech. v.22 Sequence analysis of β-D-xylosidase gene from Bacillus stearothermophilus Oh,H.J.;Y.J.Choi
  10. Kor. J. Appl. Microbiol Bioeng. v.17 Molecular cloning and expression of a xylanase gene from thermophilic alkalophilic Bacillus sp. K-17 in E. Coli Yu,J.H.;K.S.Chung;D.H.Oh
  11. J. Biol. Chem. v.193 Protein measurement with Folin phenol reagent Lowry,O.H.;N.J.Rasebrough;A.J.Farr;R.J.Randall
  12. J. Biol. Chem. v.238 Inactivation of myosin by adenosine triphosphate and other phosphate compounds Levy,H.M.;P.D.Leber;E.M.Ryan
  13. Nature(London) v.227 Cleavage of structure proteins during the assembly of the head of bacteriophage T4 Laemmili,U.K.
  14. Biochemica et Biophysica Acta v.1041 Chemical modification of xylanases Vasanti Deshpande;J.Hinge;M.Rao
  15. Biosci. Biotech. Biochem. v.57 Chemical modification of xylanases from Streptomyces sp. Mauri,M.;K.Nakanish;T.Yasui