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Ion-exchange chroma...
Ion-exchange chromatographic purification and quantitative analysis of Trichoderma reesei cellulases cellobiohydrolase I, II and endoglucanase II by fast protein liquid chromatography
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Medve, József (författare)
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Lee, Dora (författare)
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- Tjerneld, Folke (författare)
- Lund University,Lunds universitet,Biokemi och Strukturbiologi,Centrum för Molekylär Proteinvetenskap,Kemiska institutionen,Institutioner vid LTH,Lunds Tekniska Högskola,Biochemistry and Structural Biology,Center for Molecular Protein Science,Department of Chemistry,Departments at LTH,Faculty of Engineering, LTH
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(creator_code:org_t)
- 1998
- 1998
- Engelska.
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Ingår i: Journal of Chromatography A. - 0021-9673. ; 808:1-2, s. 153-165
- Relaterad länk:
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http://dx.doi.org/10...
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Abstract
Ämnesord
Stäng
- Trichoderma cellulases appear in several isoforms which makes their purification and analysis difficult. We used fast protein liquid chromatography (FPLC) to purify three major cellulases and to quantitate these enzymes in reconstituted mixtures during cellulose hydrolysis studies (in lack of specific substrates and because of the synergism between the enzymes such analysis is very difficult, if at all possible, with conventional activity measurements). For the analysis methods linear calibration was achieved from 10-15 pmol to 0.5-1 nmol (from 0.5-0.8 to 27-64 μg) for the different enzymes. Due to the high resolution chromatographic media used, our purification methods are simpler and quicker than the usual protocols for cellulase purification. Several isoforms of cellobiohydrolase (CBH) I were purified. The isoforms had different isoelectric points (pI) but their catalytic and adsorption properties were similar. A remarkable feature of CBH I and endoglucanase (EG) II was that their electrophoretically pure preparations gave double peaks during ion-exchange chromatography in certain pH intervals where the two peaks (probably representing two conformations) were transformed into each other by changing pH. This behaviour of cellulases has never been reported before and further explains the difficulties in cellulase purification.
Ämnesord
- NATURVETENSKAP -- Biologi (hsv//swe)
- NATURAL SCIENCES -- Biological Sciences (hsv//eng)
Nyckelord
- Enzymes
- Cellulases
- Cellobiohydrolases
- Endoglucanases
Publikations- och innehållstyp
- art (ämneskategori)
- ref (ämneskategori)
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