Purification and characterization of endo-β-1,4-glucanase encoded by clostridium thermocellum gene in escherichia coli = Clostridium thermocellum유전자에 의해 escherichia coli속에서 생산된 Endo-β-1,4-glucanase의 정제 및 특성

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The cellulase produced by E. coli harbouring C. thermocellum endo-$\beta$-1,4-glucanase gene was purified and characterized. Streptomycin sulfate fractionation, heat treatment, ammonium sulfate fractionation, DEAE-Sephadex A-50, and Hydroxyapatite column chromatography were employed for the purification. The purification procedures yielded a 40-fold purification of the enzyme with yield of 29\%. The molecular weight of the purified enzyme was estimated to be 40,000 dalton by SDS-polyacrylamide gel electrophoresis and gel filtration. The enzyme showed optimum activity at pH 5.0 and at $65\,^\circ\!C$. Viscometric assay showed that the enzyme behaved as an endoglucanase. The enzyme was insensitive against end product inhibition by glucose and cellobiose and resistant to organic solvents (ethanol, acetate, and lactate). The enzyme was sensitive to Mn$^{++}$, CO$^{++}$, EDTA, and Hg$^{++}$, but not to Ca$^{++}$, Mg$^{++}$, and Zn$^{++}$. Enzyme activity was stimulated by low concentrations of Cu$^{++}$, but inhibited by high concentrations of Cu$^{++}$. The values of Km and Vm of this enzyme for CMC were 0.39 \% and 268 units per mg protein, respectively. The Km an Vm for pNPC hydrolysis were 0.75 mM and 1.32 units per mg protein, respectively.
Advisors
Pack, Moo-Young박무영
Description
한국과학기술원 : 생물공학과,
Publisher
한국과학기술원
Issue Date
1989
Identifier
66672/325007 / 000871333
Language
eng
Description

학위논문(석사) - 한국과학기술원 : 생물공학과, 1989.2, [ v, 51 p. ]

Keywords

단백질 분리.

URI
http://hdl.handle.net/10203/28325
Link
http://library.kaist.ac.kr/search/detail/view.do?bibCtrlNo=66672&flag=dissertation
Appears in Collection
BS-Theses_Master(석사논문)
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