Non-hydrolytic
Disruption of Crystalline Structure of Cellulose by Cellulose Binding Domain
and Linker Sequence of Cellobiohydrolase I from Penicillium
janthinellum
GAO Pei-Ji*, CHEN Guan-Jun, WANG Tian-Hong, ZHANG
Ying-Shu, LIU Jie
( State key Laboratory of Microbial Technology, Shandong University, Jinan
250100, China )
Abstract The
cooperation between cellobiohydrolase (CBHI) and endoglucanase (EG) is
necessary for biodegradation of native cellulose, but its mechanism is still poorly
understood. The present paper report at the first time that an isolated
component, the cellulose binding domain with its linker sequence of
cellobiohydrolase I from Penicillium janthinellum (CBDCBHI),
plays an important role in the synergism between CBHI and EGI during cellulose
biodegradation. A recombinantplasmid (pUC18C), containing the gene fragment
encoding CBDCBHI from P.janthinellum was derived from
pUC18-181. In pUC 18C, the catalytic domain region of cbhI gene was
deleted by in vitro DNA manipulations and then E.coli JM 109
was transformed for the production of LacZ-CBD fusion protein. The active
LacZ-CBD fusion protein was digested by papain and then purified by
re-exclusion chromatography. The purified peptide sequence of CBDCBHI
had the ability of binding crystalline cellulose. The detailed morphological
and structural changes of cotton fibers after binding CBDCBHI were
investigated by using scanning electron microscopy, calorimetric activity and
X-ray diffraction. The results demonstrated that the CBDCBHI not
only has a high binding capacity to cellulose, but also causes non-hydrolytic
disruption of crystalline cellulose, which leads to the release of short
fibers. IR spectroscopy and X-ray diffraction show that destabilization is
caused by the non-hydrolytic disruption of cellulose and the disruption of
hydrogen bonds in crystalline cellulose. The efficiency of crystalline
cellulose degradation was enhanced by synergistic action of CBDCBHI
with EGI. These results suggest that the cellulose-binding domain with its
linker plays an important role in crystalline cellulose degradation.
Key words cellulose degradation；cellobiohydrolase I；cellulose binding domain
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