Purification and properties of Clostridium thermocellum endoglucanase 5 produced in Escherichia coli.

Abstract:

:Endoglucanase 5 (EG5) has been isolated from the strain of E. coli TG1 harboring recombinant plasmid pCU108, which contains the cel5 gene of C. thermocellum. The enzyme has been produced with 98-fold purification and a final yield of 27% by using subsequent twofold high performance ion-exchange chromatography on Mono Q and high performance chromatofocusing on Mono P. The protein has a mol mass of 35 kDa and includes 3 multiple forms with pI 4.4-4.8 as evidenced by analytical gel isoelectrofocusing. EG5 cleaves CMC (Km = 0.097 g/L, Vmax = 8.2 mg/min.mg of protein), amorphous cellulose, xylan, lichenan as a substrate with an optimum temperature of 80 degrees C and pH 6.0 and Avicel (Km = 18.2 g/L, Vmax = 0.035 mg/min.mg of protein) with an optimum temperature of 60 degrees C and pH 6.0. Cellobiose in concentrations up to 200 micrograms/mL do not inhibit the hydrolysis of CMC by EG5, but 10-30 micrograms/mL of glucose significantly decrease the activity of this enzyme. The stimulating role of calcium chloride and concentration of protein in the system has been demonstrated for Avicel hydrolysis by EG5.

authors

Mosolova TP,Kalyuzhnyi SV,Varfolomeyev SD,Velikodvorskaya GA

doi

10.1007/BF02788898

subject

Has Abstract

pub_date

1993-07-01 00:00:00

pages

9-18

issue

1

eissn

0273-2289

issn

1559-0291

journal_volume

42

pub_type

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