微生物遺伝資源利用マニュアル(16)

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1 (16) MAFF Microorganism enetic Resources Manual No.16 Enzymes of filamentous fungi in fermentation Aspergillus oryzae (1) α-ec β-(EC )-1,4-α-(EC )α- EC α-1,6 EC α-1,6 α-1,4 EC

2 (A) (B) (C) α- α-1,4 β- Bacillus polymyxa, Bacillus megaterium β- α- β- α-1,4 α-1,6 α- α- α-α-1,4 endo α-1,6 α- α-taka-amylase A ) ) 2 53,662 Ca 2+ Ca 2+ 2

3 PDBID: 2TAA, exo α-1,6 (2) α- endo β exo Somogy-Nelson 1% Somogy Nelson 12mm 200 µl 50 µl Somogy 250 µl 20 Nelson 500 µl ml 500 nm 3 3

4 500nm (1) (EC ), 1,4-β-EC , β-ec Aspergillus awamori, Aspergillus usami Aspergillus oryzae 4

5 β-1, ) 4) S1S2S3 35% 4050% 90% 5) Acetobacter xylinum β-1,4 6,00010,000 8,00010,000 Valonia 44,000 6) D- 180 C3-OHO C6-OHC2-OH nm 7) X X 65% 70% 8) 1 4 5

6 6

7 Butyrivibrio fibrisolven s Cellulomonas fimi Cellulomonas uda Cellvibrio gilvus Clostridium thermocellum Pseudomonas fluorescence Ruminococcus albus Streptomyces lividans Thermomonospora curvata Thermomonospora fusca Aspergillus aculeatus Aspergillis niger Aspergillus oryzae Chaetomium cellulolyticum Humicola insolens Irpex lacteus Phanerochaete chrysosporium Penicillium purpurogenum Schizophyllum commune Sporotrichum thermophile Trichoderma reesei Trichoderma viride Trichoderma koninngi 9) O-EC X (EC ) β- (EC ) 1,4-β- (EC ) 1,4-β- (EC ) EC CMC CMC CMC -1,4-β- 1,4-β-EC ,4-β-EC ,4-β- CMC β-ec ,4-β- 1,4-β- β- 7

8 β Reese 10) C1-Cx C1-CxC1 Cxβ- Wood 11) Trichoderma koningiic1 1,4-β-D-CBH Trichoderma viride CBH 12) T. virideβ )CBH 15, 16) β- 15, 17) C1 Cx OkadaT. viri de 18)12 1 β ) MuraoAspergillus aculeatus 22) 9 Aspregillus 5) A. aculeatus 5) 1906 Seillère 1912 Pringsheim 1950 Reese C1-Cx 1951 Siu hydrogen bondase 1953 Whitaker 1953 Marsh 1968 Okada Trichoderma viride 1972 Wood Trichoderma koningii 1979 Wood Kanda Irpex lacteus 1983 Shoemaker Trichoderma reesei 1983 Béguin Clostridium thermocellum 1983 Lamed C. thermocellum 1987 Murao Aspergillus aculeatus 1988 Okada T.viride 8

9 Clostridium thermocellum 23) C. thermocellum 24) Aspergillus oryzaetrichoderma reeseiclostridium thermocellum CBH 25) β ) HCA 29) (1) α-(2) α-(3) HCA glycosyl hydrolase family (HF)HF β- 3 ( (2) Cellulose Azure (Sigma) Cellulose AzureFernley 30) Remazol Brilliant Blue R 0.51% Cellulose Azure Czapeck-Dox 57 9

31-32) 50 g 60% H2SO4 200 ml1 2 L No.3 0.5 L 300 ml 500 ml 1N NaOHpH 56 ph 75% 3 L 99.

10 31-32) 50 g 60% H2SO4 200 ml1 2 L No L 300 ml 500 ml 1N NaOHpH 56 ph 75% 3 L 99.5% 12% Czapeck-Dox 57 CMCCMC D.S., degree of substitution CMCSomogy-Nelson CMCCMCDinitro salicylic acid (DNS) 33) DNSSomogy-Nelson 1/10 CMC 10

11 low viscositysigma Dinitro salicylic acid (DNS) A 4.5% NaOH 300 ml 1% DNS 880 ml 225 g B10% NaOH 22 ml 10 g 100 ml 69 ml NaHCO3 6.9 g B A 2 1% CMC 0.1 ml DNS 0.3 ml ml 500 nm 11

12 (1) N C (EC ) (EC )(EC )(EC ) (EC ) ( Aspergillus sojae Aspergillus oryzae (2) 34) 2% 0.1M ph 7 0.4M 4 ml 1 ml mL 3030 No nm No. 5 1 ml 0.4M 5 ml 2 1 ml nm 12

13 13

14 1) (1983) 2) Y. Matsuura et al.; Molecular structure of taka-amylase A. I. Backbone chain folding at 3 A resolution., J. Biochem (Tokyo), 87, (1980) 3) pp.98 (1981) 4) R.L. Whistler and E.L. Richards; Hemicellulose, The Carbohydrates IIA, Ed. by W. Pigman and D. Horton, pp.448, Academic Press, NY (1970) 5) (1987) 6). Franz and W. Blaschek; Cellulose., Methods in Plant Biochemistry, 2, Ed. by P. M. Day, pp.301, Academic Press, NY (1990) 7). Franz and W. Blaschek; Cellulose., Methods in Plant Biochemistry, 2, Ed. by P. M. Day, pp , Academic Press, NY (1990) 8) D. Fegnel and. Wegener, Wood; Chemistry, Ultrastructure, Reactions, Walter de ruyter, Berlin and NY (1984) 9) Enzyme nomenclature, Recommendation of the Nomenclature Committee of the International Union of Biochemistry and Molecular Biology, Academic Press, N.Y.(1992) 10) E.T. Reese, R..H. Siu and H. S. Levinson; The biological degradation of soluble cellulose derivatives and its relationships to the mechanism of cellulose hydrolysis., J. Bacteriol., 59, (1950) 11) T.M. Wood and S.I. Maccrae; The purification and properties of the C1 component of Trichoderm a koningii cellulase., Biochem. J., 128, (1972) 12) L.E.R. Berghem and L.. Pettersson; The mechanism of enzymatic cellulose degradation Purification of a cellulolytic enzyme from Trichoderma viride., Eur. J. Biochem., 37, (1973) 13) L.E.R. Berghem, L.. Pettersson and U. -B. Axio- Fredriksson; The mechanism of enzymatic cellulose degradation- Purification and some properties of 2 different 1,4-β-glucan glucanohydrolases from Trichoderma viride., Eur. J. Biochem., 61, (1976) 14) S.P. Shoemaker and R.D. Brown Jr.; Characterization of endo 1,4-β-D-glucanase purified from Trichoderma viride., Biochim. Biophys. Acta, 523, (1978) 15). Beldman, M. F. Searle-Van Leeuwen, F.M. Rombouts and F..J. Voregen; The cellulase of Trichoderma viride - Characterization and comparison of all detectable endoglucanases, exoglucanases and b-glucosidases., Eur. J. Biochem., 146, (1985) 14

15 16) E.K. um Jr. and R.D. Brown Jr.; Structural characterization of glycoprotein cellulase 1,4-β-D-glucan cellobiohydrolase C from Trichoderma viride., Biochim. Biophys. Acta, 446, (1976) 17) L.E.R. Berghem and L.. Pettersson; The mechanism of cellulose degradation- Isolation and some properties of a β- glucosidase from Trichoderma viride., Eur. J. Biochem., 46, (1974) 18) 35, (1988) 19). Okada; Enzymatic studies on a cellulase system of Trichoderma viride Part 2 Purification properties of 2 cellulases., J. Biochem., 77, (1975) 20). Okada and K. Nishizawa; Enzymatic studies on a cellulase system of Trichoderma viride Part 3 Trans glycosilation properties of 2 cellulase components of random type., J. Biochem., 78, (1975) 21). Okada; Enzymatic studies on a cellulase system of Trichoderma viride Part 4 Purification and properties of a less random type cellulase., J. Biochem., 80, (1976) 22) S. Murao and J. Nakamoto; Isolation and identification of a cellulolytic enzyme producing microorganism., J. Fermrnt. Technol., (1979) 23) J. Petre, R. Rongin and J. Millet; Purification of an endo- 1,4-β-glucanase from Clostridium thermocellum., Biochimie, 63, (1981) 24) R. Lamed, E. Setter and E.A. Bayer; Characterization of a cellulose binding, cellulase conteining complex in Clostridium thermocellum., J. Bacteriol., 156, (1983) 25) S. Shoemaker, V. Schweickart, M. Lander, D. elfand, S. Kwok, K. Myambo and M. Innis; Molecular cloning of exo-cellobiohydrolase I derived from Trichodeima reesei strain L27., BIO/TECHNOLOY, 1, (1983) 26) P. Beguin, M. Rocancourt, M.C. Chebrou, and J.P. Aubert; Mapping of mrna encoding endoglucanase A from Clostridium thermocellum., Mol. en. enet., 202, (1986) 27) O. repinet and P. Beguin; Sequence of cellulase gene of Clostridium thrmocellum coding endoglucanase B., Nucleic Acid Res. 14, (1986) 28) D. Petre, J. Millet, R. Rongin, P. Beguin, H. irard and J.P. Aubert; Purification and properties of the endoglucanase C of Clostridium thermocellum produced in Escherichia coli., Biochimie, 68, (1986) 29) C. abouriaud. V. Bissery, T. Benchetrit and J.P. Mornon; Hydrophobic cluster analysis: an efficient new way to compare and analysis amino acid sequences., FEBS Lett., 224, (1987) 15

16 30) H.N. Fernley; The use of reactive dyestuffs in enzymology: new substrates for cellulolytic enzymes., Biochem J., 87, 90-5 (1965). 31) N.R. ilkes, B. Henrissat, D.. Kilburn, R.C. Miller Jr. and A.J. Warren; Domains in microbial β-glucanase; sequence conservation, function, and enzyme families., Microbiol. Rev., 55, (1991) 31)., ;.,26, (1979) 32) T. Sasaki, T. Tanaka, N. Nanbu, Y. Sato and K. Kainuma; Correlation between X-ray diffraction measurements of cellulose crystalline structure and the susceptibility to microbial cellulase., Biotechnol. Boieng., 21, (1979) 33).L. Miller; Use of dinitrosalicylic acid reagent for determination of reducing sugar., Anal. Chem., 31, (1959) 34) ;9, (1983) 16 2 January, National Institute of Agrobiological Sciences

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untitled CH 2 H H H CH 2 n-1 H CH 2 n H Potato α-glucan phosphorylase H n soluble starch P i n-1 H P() 2 glucose-1-phosphate Pi H CBPase H glucose P i glucose-1-phospate cellobiose H cellooligosaccharide n+1 CDPase