Production of Glucoamylase from Hybrid Constructed by Intergenic Nuclear Transfer between Saccharomycopsis sp. and Saccharomyces sp.

핵전이법에 의해 형성된 Saccharomycopsis 속과 Saccharomyces 속의 잡종에서 glucoamylase 생산에 관한 연구

  • 양영기 (조선대학교 자연과학대학 생물과학부) ;
  • 임채영 (조선대학교 자연과학대학 생물과학부) ;
  • 김종권 (조선대학교 자연과학대학 생물과학부) ;
  • 문명님 (조선대학교 자연과학대학 생물과학부) ;
  • 이영하 (충남대학교 미생물학과)
  • Published : 2001.09.01

Abstract

The glucoamylase was purified from the induced culture filtrate of hybrid between Saccharomycopsis sp. and Saccharomycopsis sp. made by nuclear transfer and characterized for some enzyme properties. The enzymewas purified 76-fold in an overall yield of 16% from the culture medium by ammonium sulfate fractionation,Sephadex G-150 gel permeation chromatography and DEAE-Sephadex A-50 ion exchage chromatography.The molecular weight of the purified glucoamylase was estimated to be 57.5 KDa on SDS-polyacrylamidegel electrophoresis and Sephadex G-150 gel permeation chromatography. The purified enzyme was active atpH-5.0 and $40^{\circ}C$. The Km value for soluble starch was 2.6 mg/ml. The enzymatic activity was stimulated inthe presence of TEX>$Ca^{2+}$, EDTA, $Co^{2+}$, $Mg^{2+}$, and $Mn^{2+}$

핵전이에 의해 형성된 잡종의 유도 배양액으로부터 glucoamylase를 정제하고 몇몇의 효소 특성을 조사하였다. 효소는 76배 정제하였고, ammonium sulfate fractionation, Sephadex G-150 gel permeation chromatography 그리고 DEAE-Sephadex A-50 ion exchange chromatography의 순서 배양액으로부터 정제한 결과 전반적인 수율은 16%를 나타내었다. SDS-polyacrylamide gel electrophoresis와 Sephadex G-150 gel permeation chromatography에 의하여 정제된 glucoamylase의 분자량을 측정한 결과 57.5 kDa를 나타내었으며, 정제된 효소의 최적 pH와 온도는 각각 5.0과 $40^{\circ}C$로 나타났다. 또한 가용성 전분에 대한 Km값은 2.6 mg/ml을 나타내었으며, 정제된 효소는 $Ca^{2+}$, EDTA, $Co^{2+}$, $Mg^{2+}$, 그리고 $Mn^{2+}$의 존재하에 활성이 중진됨을 알수 있었다.

Keywords

References

  1. Microbios v.80 Purification and characterization of the glucoamylase produced by a strain of Aspergillus flavus Abyad, M.S.;A.R. Shanshoury;M. Hafez
  2. Folia Microbiol. v.41 Purification and properties of two forms of glucoamylase from Aspergillus niger Amirul, A.A.;S.L. Khoo;M.N. Nazalan;M.S. Razip;M.N. Azizan
  3. Folia Microbiol v.46 Thermostable glucose-tolerant glucoamylase produced by the thermophilic fungus Scytalidium thermophilum Aquino, A.C.;J.A. Jorge;H.F. Terenzi;M.L. Polizeli
  4. Folia Microbiol v.45 Polysaccharide hydrolases of Aureobasidium pullulans Augustin, J.
  5. Biotech. Bioeng. v.76 Optimization of maltodextrin hydrolysis by glucoamylase in a batch reactor Cepeda, E.;M. Hermosa;A. Ballesteros
  6. Biochem. J. v.349 H-dependence of the fast step of maltose hydrolysis catalysed by glucoamylase G1 from Aspergillus niger Christensen. U.
  7. J. Chromatogr. B. Biomed. Sci. Appl. v.743 Bioaffinity extraction of glucoamylase in aqueous two-phase systems using starch as free bioligand De Gouveia, T.;B.V. Kilikian
  8. Microbiol v.50 Purification and characterization of extracellular amylolytic enzymes from the yeast viron De Mot, R.;H. Verachtert
  9. FEBS Lett v.471 Evidence that the glucoamylases and alpha-amylase secreted by Aspergillus niger are proteolytically processed products of a precursor enzyme Dubey, A.K.;C. Suresh;R. Kavitha;N.G. Karanth;S. Umesh-Kumar
  10. J AOAC lnt. v.78 Glucoamylase activity in industrial enzyme preparations using colorimetric enzymatic method: collaborative study Elder, M.T.;R.S. Montgomery
  11. J. Gen. Microbiol v.136 Comparison of two glucoamylases from Hormoconis resinae Fagerstrom, R.;A. Vainio;K. Suoranta;T. Pakula;N. Kalkkinen;H. Torkkeli
  12. Biosci. Biotechnol. Biochem. v.62 Purification and properties of an amylopullulanase, a glucoamylase, and an alpha-glucosidase in the amylolytic enzyme system of Thermoanaerobacterium thermosaccharolyticum Ganghofher D.;J. Kellermann;W.L. Staudenbauer;K. Bronnenmeier
  13. J. Biotechnol v.67 Coimmobilization of glucoamylase and glucose isomerase by molecular deposition technique for one-step conversion of dextrin to fructose Ge Y.;Y. Wang;H. Zhou;S. Wang;Y. Tong;W. Li
  14. FEMS Microbiol. Lett. v.54 Characterization of glucoamylase from Aspergillus terreus 4 Ghose, A.;B.S. Chatterjee;A. Das.
  15. J. Appl. Bacteriol. v.71 Purification and chraterization of glucoamylase of .Aspergillus terreus NA- 17O mutant Ghosh, A.;B. Chatterjee;A. Das
  16. Appl. Biochem. Biotechnol. v.86 Study of biocatalyst to produce ethanol from starch. Coimmobilization of glucoamylase and yeast in gel Giordano, R.L.;P.C. Hirano;L.R. Goncalves;W.S. Netto
  17. Biosci. Biotechnol. Biochem. v.59 Role of the carbohydrate moiety of a glucoamylase from Aspergilus awamori var. Kawachi in the digestion of raw starch Goto, M.;E. Kuwano;W. Kanlayakrit;S. Hayashida
  18. J. Appl. Bacteriol. v.79 Cultural conditions for production of glucoamylase from Lactobacillus amylovorus ATCC 33621 James, J.A.;B.H. Lee
  19. Curr. Microbiol. v.34 Purification of glucoamylase from Lactobacillus amylovorus ATCC 33621 James J.A.;J.L. Berger;B.H. Lee
  20. Natrue v.227 Clevage of structural proteins during the assembly of the head of bacteriophage T4 Laemmli, U.K.
  21. J. Appl. Biochem v.5 Fungal glucoamylaseseng Manjunath, P.;B.C. Shenoy;MR. Raghavendra Rao
  22. J. Chromatogr. B. Biomed. Sci. Appl. v.711 Separation and purification of glucoamylase in aqueous two-phase systems by a two-step extraction Minami, N.M.;B.V. Kilikian
  23. Appl. Microbiol. Biotechnol. v.50 Purification and propertics of the raw-starch-digesting glucoamylases from Corticium rolfsii Nagasaka. Y.;K. Kurosawa;A. Yokota;F. Tomita
  24. Biochemistry v.35 Identification of enzymesubstrate and enzyme-product complexes in the catalytic mechanism of glucoamylase from asperillus awamori Natarajan, S.K.;M.R. Sierks
  25. Biotechnol. Appl. Biochem Advances in microbial amylases Pandey, A.;P. Nigam;C.R. Soccol;V.T. Soccol;D. Singh;R. Mohan
  26. Kor. J. Appl. Microbiol. Bioeng. v.17 Some factors affecting glucoamulase production from Aspergillus sp. Park, I.S.;Y.H. Chung
  27. Appl. Microbiol. Biotechnol. v.53 Glucoamylase production in batch, chemostat and fed-batch cultivations by an industrial stranin of Aspergillus niger Pedersen, H.;M. Beyer;J. Nielsen
  28. J. Microbiol. Methods v.42 Relating growth dynamics and glucoamylase excretion of individual Saccharomyces cerevisiae cells Porro, D.;M. Venturini;L. Brambilla;L. Alberghina;M. Vanoni
  29. Biochemistry v.40 Stability and function of interdomain linker variants of glucoamylase 1 from Aspergillus niger Sauer J.;T. Christensen;T.P. Frandsen;E. Mirgorodskaya;K.A. McGuire;H. Driguez;P. Roepstorff;B.W. Sigurskjold;B. Svensson
  30. Lett. Appl. Microbiol. v.23 Purification and characterization of glucoamylase produced by Aspergillus niger in solid state fermentation Selvakumar P.;L. Ashakumary;A. Helen;A. Pandey
  31. Biotechnol. Prog. v.13 Solvent and viscosity effects on the rate-limiting product release step of glucoamylase during maltose hydrolysis Sierks M.R.;C. Sico;M. Zaw
  32. J. Inst. Brew. v.90 Isolation and characterization of the amylolytic system of Schwanniomyces castellii Sills, A.M.;M.E. Sauder;G.G. Stewart
  33. J. Basic Microbiol. v.35 Simultaneous raw starch hydrolysis and ethanol fermentation by glucoamylase from Rhizoctonia solani and Saccharomyces cerevisiae Singh, D.;J.S. Dahiya;P. Nigam
  34. FEBS Lett. v.447 The starch-binding domain from glucoamylase disrupts the srtucture of starch Southall, S. M.;P. J. Simpson;H. J. Gilbert;G. Willialnson;M. P. Williamson
  35. Electrophoresis v.20 Separation and direct detection of raw and gelatinized starch hydrolyzing activities of glucoamylase on isoelectric focusing gels Suresh C.;A. K. Dubey;R. Kini;S, Umesh-Kumar;N. G. Karanth
  36. J. Biochem. v.84 Purification and some properties of three forms of glucoamylase from a Rhizopus species Takahashi, T.;Y. Tsuchida;M. Irie
  37. Biosci. Biotechnol. Biochem. v.62 Thermal unfolding of the starch binding domain of Aspergillus niger glucoamylase Tanaka A.;S. Karita;Y. Kosuge;K. Senoo;H. Obata;N. Kitamoto
  38. Enzyme Microbiol. Technol. v.5 Behaviour of Endomycopsis fiburigera glucoamylase towards raw starch Ueda, S.;B.C. Saba
  39. Biochim Biophys. Acta. v.1527 The effect of pH on glucoamylase production, glycosylation and chemostat evolution of Aspergillus niger Wallis, G.L.;R.J. Swift;R. Atterbury;S. Trappe;U. Rinas;F.W. Hemming;M.G. Wiebe;A.P. Trinci;J.F. Peberdy
  40. Appl. Environ. Microiol. v.44 Isolation and characterization of Schwanniomyces alluvius amylolytic enzymes Wilson, J.J.;W.M. Ingledew
  41. Handbook of amylase and related enzymes, their sources isolation methods, properties and applications Yammamoto, T.
  42. Agric. Biol. Chem. v.49 Purification and properties of an amylase from Bacillus cereus NY- 14 Yoshigi, N;J. Chikano;M. Kamimura