|Skory, Christopher - Chris|
Submitted to: Biotechnology for Fuels and Chemicals Symposium Proceedings
Publication Type: Abstract Only
Publication Acceptance Date: 5/1/2002
Publication Date: 5/1/2002
Citation: SUN, Y., ADNEY, W.S., BERGMANN, B.A., CHENG, J., DECKER, S.R., FREER, S.N., HIMMEL, M.E., NISHIMURA, Y., SKORY, C.D., TISSERAT, B. EXPRESSION OF ENDOGLUCANASE E1 IN TRANSGENIC DUCKWEED LEMNA MINOR. BIOTECHNOLOGY FOR FUELS AND CHEMICALS SYMPOSIUM. 2002. ABSTRACT NO. 6-42. Interpretive Summary:
Technical Abstract: Transgenic duckweed (Lemna minor) that expresses Acidothermus cellulolyticus E1 endoglucanase was generated using Agrobacterium-mediated transformation. Out of 15 independent transgenic lines, 1 line with the highest CMC-degrading activity was selected for further studies. The 2-week-old transgenic duckweed fronds cultured in SH medium were ground in sodium citrate buffer (pH 4.8), and the expression of endoglucanase E1 in the supernatant was examined using immunoblotting and enzyme assays. The E1 produced in transgenic duckweed was active, and the expression level was 0.24% of total soluble protein. The recombinant E1 protein was thermostable and had higher activity at 70 deg C compared to room temperature. The average E1 activity at 70 deg C in duckweed extracts was 0.21 umol min-1 g-1 fresh duckweed or 0.17 umol min-1 mg-1 soluble protein. A 5-min heat treatment of the extract at 65 deg C effectively removed most other proteins without reducing the enzyme activity. This may be useful for downstream processing.