TY - JOUR
T1 - Monoclonal antibodies against core and cellulose-binding domains of Trichoderma reesei cellobiohydrolases I and II and endoglucanase I
AU - Aho, Sirpa
AU - Olkkonen, Vesa
AU - Jalava, Taina
AU - Paloheimo, Marja
AU - Buhler, Rolf
AU - Niku-Paavola, Marja-Leena
AU - Bamford, Dennis
AU - Korhola, Matti
N1 - Project code: BIO9003
PY - 1991
Y1 - 1991
N2 - Cellulases from Trichoderma reesei form an enzyme
group with a common structural organization. Each cellulase enzyme is
composed of two functional domains, the core region containing the
active site and the cellulose‐binding domain (CBD). To facilitate the
specific detection of each domain, monoclonal antibodies (mAb) against
cellobiohydrolase I (CBHI), cellobiohydrolase II (CBHII) and
endoglucanase I (EGI) were produced. Five mAb were obtained against
CBHI, ten against CBHII and eight against EGI. The location of the
antigenic epitope for each antibody was mapped by allowing the
antibodies to react with truncated cellulases, synthesized from deleted
cDNA in Saccharomyces cerevisiae. Proteolytic fragments of Trichoderma
cellulases, obtained by papain digestion, were used to confirm the
results. Specific antibodies were detected against the core and the CBD
epitopes for all three cellulases. Using the truncated enzymes, it was
possible to locate the epitopes to a reasonably short region within the
protein. To obtain a quantitative assay for each enzyme, a specific mAb
against each antigen was chosen, based on the affinity to the
corresponding antigen on Western‐blot staining and on filter blots of
the cellulolytic yeasts. The mAb were used to quantitative the
corresponding enzymes in T. reesei culture medium. Specific
quantitation of each cellulase enzyme has not been possible by
biochemical assays or using polyclonal antibodies, due to their
cross‐reactions. Now, these mAb can be specifically used to recognize
and quantitate different domains of these three important cellulolytic
enzymes.
AB - Cellulases from Trichoderma reesei form an enzyme
group with a common structural organization. Each cellulase enzyme is
composed of two functional domains, the core region containing the
active site and the cellulose‐binding domain (CBD). To facilitate the
specific detection of each domain, monoclonal antibodies (mAb) against
cellobiohydrolase I (CBHI), cellobiohydrolase II (CBHII) and
endoglucanase I (EGI) were produced. Five mAb were obtained against
CBHI, ten against CBHII and eight against EGI. The location of the
antigenic epitope for each antibody was mapped by allowing the
antibodies to react with truncated cellulases, synthesized from deleted
cDNA in Saccharomyces cerevisiae. Proteolytic fragments of Trichoderma
cellulases, obtained by papain digestion, were used to confirm the
results. Specific antibodies were detected against the core and the CBD
epitopes for all three cellulases. Using the truncated enzymes, it was
possible to locate the epitopes to a reasonably short region within the
protein. To obtain a quantitative assay for each enzyme, a specific mAb
against each antigen was chosen, based on the affinity to the
corresponding antigen on Western‐blot staining and on filter blots of
the cellulolytic yeasts. The mAb were used to quantitative the
corresponding enzymes in T. reesei culture medium. Specific
quantitation of each cellulase enzyme has not been possible by
biochemical assays or using polyclonal antibodies, due to their
cross‐reactions. Now, these mAb can be specifically used to recognize
and quantitate different domains of these three important cellulolytic
enzymes.
U2 - 10.1111/j.1432-1033.1991.tb16227.x
DO - 10.1111/j.1432-1033.1991.tb16227.x
M3 - Article
VL - 200
SP - 643
EP - 649
JO - FEBS Journal
JF - FEBS Journal
SN - 1742-464X
IS - 3
ER -