Putative Cellulase Research Articles

Cel1, probably encoding a cellobiohydrolase lacking the substrate binding domain, is expressed in the initial infection phase of Claviceps purpurea on Secale cereale.

At the host-pathogen interface of hyphae penetrating host cell walls in the rye ovary, a lack of cellulase-gold labeling of beta-1, 4-glucan in host cell walls indicates that enzymatic degradation of cellulose might be an important factor during the infection of rye by Claviceps purpurea. Using cbh1 from Trichoderma reesei as a probe, a putative cellulase gene (cel1) was isolated from a genomic library of the C. purpurea strain T5. The coding region of 1,616 bp contains two introns and a putative signal peptidase cleavage site, leaving a coding capacity of 437 amino acids for the mature protein. The derived amino acid sequence shares significant homology with other fungal cellobiohydrolases and lacks the substrate binding domain. Expression analysis using reverse transcriptase-polymerase chain reaction (RT-PCR) shows that cel1 is induced during the first days of infection of rye by C. purpurea. It may be involved in the penetration and degradation of host cell walls by depolymerizing plant beta-1, 4-glucan and, therefore, play a role in the infection process.

Nucleotide sequences of the celB gene encoding endo-1,4-β-β-glucanase-2, ORF1 and ORF2 forming a putative cellulase gene cluster of Clostridium josui

Abstract The nucleotide sequence of the ce1B gene coding for endo-1,4-β-glucanase-2 (EG-2) of Clostridium josui was determined. The structural gene consists of an open reading frame of 1,380 by encoding a 460-amino acid polypeptide. The deduced amino acid sequence of EG-2 shows high similarities with the Clostridium cellulolyticum CelCCC (92.3%) and the Clostridium thermocellum endoglucanase CelA (61.6%) in the catalytic domain, but no homology with EG-1 of C. josui . In the COOH-terminal region of EG-2, there exists a reiterated segment similar to the segments present in many clostridial cellulases and hemicellulases. Upstream and downstream of the celB gene, two incomplete open reading frames (ORF1 and ORF2) were found to encode proteins homologous to the proteins which consist of the C. cellulolyticum cellulase gene cluster. ORF1 containing a reiterated segment showed high similarities with the proteins deduced from each ORF1 from C. cellulolyticum and Caldocellum saccharolyticum . Downstream of the ce1B gene, ORF2 was found to encode a protein homologous to CeICCG of C. cellulolyticum and CelF of C. thermocellum . These results indicate that the celB gene, ORF1 and ORF2 formed a cellulase gene cluster strictly homologous to that of C. cellulolyticum , which is proposed to be part of a single operon.

CHANGES IN RIPENING ENZYMES DURING SWEET CHERRY FRUIT DEVELOPMENT

The activities of the fruit ripening enzymes cellulase, polygalacturonase (PG) and pectin methylesterase (PME) were detected during the development of sweet cherry (Prunus avium L.) fruit. Cellulase and PG activities of pericarp tissue increased 4-10 times between hypanthium abscission and harvest. PME activity remained high throughout this period of fruit development. There was a positive correlation between the anthocyanin content of the pericarp and both cellulase and PG activities. Concomitant with the increases in the activities of these ripening enzymes was a decrease in fruit firmness. The increases in cellulase and PG activities were checked following two-weeks storage at 10 C after harvest. The purification and characterization of the putative cellulase and PG enzymes will be discussed, together with attempts to chemically inhibit their activities and modify fruit softening.