EXPLORATION OF BROTH CHICKEN GUT AS GROWTH MEDIA OF Escherichia coli BL21 pET-Endo FOR Endo-β-1,4-D-Xylanase PRODUCTION

Anak Agung Istri Ratnadewi, Moch. Yoris Alidion, Agung Budi Santoso, Ika Oktavianawati

Abstract

Endo-β-1,4-D-xylanase is a hydrolytic enzyme that breakdown the 1.4 chain of xylan polysaccharide. We have succes to transform the plasmid pET-Endo gene encoding endo-1,4-β-D-xylanase from Bacillus sp. Originaly from termites abdominal to E. coli BL21. The clone was ready for large scale of enzyme production. To reduce production cost we look for subtitute media for the expensive Luria Berthani broth. Chicken guts broth is good alternative while rich of protein and very cheap. Growth of E. Coli BL21 in Chicken guts broth and LB broth (as control) was observed by Optical Density (OD) with spectrofotometer. Concentration of glucose added in broth and incubation temperature was varied. The result showed that optimal growth was in addition of 1,5 % glucose and incubated at  37 oC for 16 hours. This optimal condition was used  for E. coli BL21 pET-Endo for xylanase production. Enzyme purification have done by Ni-NTA affinity chromatography. Highest protein yield was 0,076 mg/mL obtained in 100 mM imidazole elucidation. Xylanase characteization were : activity 0,042 U/mL, specific activity      0,556 U/μg, purification factor 3,16 times and molecular weight ± 30.000 Dalton. Compare to cultivation in LB media, the enzyme activity is lower also the purity. Further optimization is neededfor higher activity and purity of the enzyme. 

Keywords

Endo-β-1,4-D-xylanase; E. Coli BL21; broth of chicken gut; glucose

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