Penicillium Canescens Case Study

Wolkov et al., (2014) cited Penicillium canescens is a filamentous fungus that typically does not secrete sufficient levels of cellulase activity. Cellobiohydrolase I (CBH) of P. canescens (PcCel7A) homologously cloned and then expressed into a host strain RN3-11-7 (niaD-). The recombinant enzyme showed maximum activity at pH 4.0–4.5. They found that PcCel7A was stable at 50 °C and pH 4.5 for 3 hours, while at 60°C after 30 min of incubation it lost 45% of activity. The recombinant enzyme showed a higher performance in prolonged hydrolysis of Avicel and milled aspen wood than CBH I (Cel7A) when equalized by protein concentration from Trichoderma reesei. Pavel reported that PcCel7A was the most industrially utilized cellulase and due to its
In this research, they used fungal host to express Cel7A enzymes from different fungi and performed assays for thermostability. They used Cel7A of Hypocrea jecorina as a reference. They found most stable homologues, Humicola grisea var. thermoidea Cel7A exhibits a 10ºC higher melting temperature (Tm of 72.5ºC) which is 4–5 times higher hydrolysis rate than H. jecorina Cel7A on phosphoric acid-swollen cellulose. The enzyme shows 57% sequence identity with H. jecorina Cel7A consisting catalytic module of GH7. The crystal structure of the H. grisea var. thermoidea Cel7A catalytic module (1.8 A ° resolution; Rwork and Rfree of 0.16 and 0.21, respectively) was similar to CBHs of other
Pectobacterium carotovorum subsp. carotovorum causes soft rot by maceration of pant cell wall. It has ability to secret hydrolytic enzymes like polygalacturonases and cellulases to destroy plants. Amplification of peh, celB and celC were performed by using P. carotovorum ATCC™ no. 15359 as a source of DNA. Primer designing and amplification of genes sequences from DNA of P. carotovorum were also performed. The pTAC-MAT-2 expression vector was used to clone the individual PCR products which then transformed into E. coli. Catalytically active domains were studied by analyzing deduced amino acid sequences of genes. SDS-PAGE analysis was carried out to estimate the molecular weights of proteins. Molecular weights of peh, celB and celC products were found to be 41.5 kDa, 29.5 kDa and 40 kDa, respectively. Agar diffusion assays were performed to qualitatively determine the activities of polygalacturonase and cellulase of the cloned