The use of ion exchange chromatography of DEAE-sepharose CL-6B in the puricication of cellulase from cellulomonas CSI-17
Purwadaria, M.B. Tresnawati
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Ion exchange chromatography of DE.AE-sepharose CL-6B was used to purify cellulase components from a culture filtrate of Ceiiuiomonas CSI-17 grown on microcrystalline cellulose Sigmaeell-20. The cellulase components were separated into three activity peaks after elution with a linear gradient of NaCl concentrations from 0 to 0.SM. The first and second peaks I. and t contained high activity for degradation of amorphous cellulose CMCase, while the third peak I, contained activity for degradation of microcrystalline cellulose avieclase. In the higher concentration of NaCI 0.0 to 1.0 M and in the same linear gradient addition the filtrate was separated into four peaks. Three peaks of 1t, IIb and It, had the same activity as l, t, and I,, while the last peak It4 had high CMCaae activity. In this separation the resolution between t1 and It, was poorer than I, and 1b* Therefore, in the further experiment the separation was carried out with stepwise gradients ofNaCI 0.08, 0.18,0.30 and 1.0 M. The choice of salt concentrations was based on results of the second linear gradients system. Five peaks were resolved well after the elution. Further enzyme determination shows that 11b has high CMCase activity, II, has high avicelase aetivity,while II. has cellobiohydrolaseactivity.