Genome-wide characterization of cellulases from the hemi-biotrophic plant pathogen, Bipolaris sorokiniana, reveals the presence of a highly stable GH7 endoglucanase
| Autor: | Pritha Kundu, Ravi K. Singh, Shree P. Pandey, Shritama Aich, Supratim Datta |
|---|---|
| Jazyk: | angličtina |
| Rok vydání: | 2017 |
| Předmět: |
0301 basic medicine
lcsh:Biotechnology Cell wall-degrading enzymes Cellulase Management Monitoring Policy and Law Applied Microbiology and Biotechnology Genome lcsh:Fuel Microbiology Pichia pastoris 03 medical and health sciences GH7 endoglucanases lcsh:TP315-360 lcsh:TP248.13-248.65 Glycoside hydrolase Alkaliphilic Gene Thermostable chemistry.chemical_classification biology Renewable Energy Sustainability and the Environment Research Glycosyl hydrolase Bipolaris biology.organism_classification Enzyme assay Ionic liquids Salt tolerant 030104 developmental biology General Energy Enzyme Biochemistry chemistry biology.protein Biotechnology Bipolaris sorokiniana |
| Zdroj: | Biotechnology for Biofuels, Vol 10, Iss 1, Pp 1-14 (2017) Biotechnology for Biofuels |
| ISSN: | 1754-6834 |
| DOI: | 10.1186/s13068-017-0822-0 |
| Popis: | Background Bipolaris sorokiniana is a filamentous fungus that causes spot blotch disease in cereals like wheat and has severe economic consequences. However, information on the identities and role of the cell wall-degrading enzymes (CWDE) in B. sorokiniana is very limited. Several fungi produce CWDE like glycosyl hydrolases (GHs) that help in host cell invasion. To understand the role of these CWDE in B. sorokiniana, the first step is to identify and annotate all possible genes of the GH families like GH3, GH6, GH7, GH45 and AA9 and then characterize them biochemically. Results We confirmed and annotated the homologs of GH3, GH6, GH7, GH45 and AA9 enzymes in the B. sorokiniana genome using the sequence and domain features of these families. Quantitative real-time PCR analyses of these homologs revealed that the transcripts of the BsGH7-3 (3rd homolog of the GH 7 family in B. sorokiniana) were most abundant. BsGH7-3, the gene of BsGH7-3, was thus cloned into pPICZαC Pichia pastoris vector and expressed in X33 P. pastoris host to be characterized. BsGH7-3 enzyme showed a temperature optimum of 60 °C and a pHopt of 8.1. BsGH7-3 was identified to be an endoglucanase based on its broad substrate specificity and structural comparisons with other such endoglucanases. BsGH7-3 has a very long half-life and retains 100% activity even in the presence of 4 M NaCl, 4 M KCl and 20% (v/v) ionic liquids. The enzyme activity is stimulated up to fivefold in the presence of Mn+2 and Fe+2 without any deleterious effects on enzyme thermostability. Conclusions Here we reanalysed the B. sorokiniana genome and selected one GH7 enzyme for further characterization. The present work demonstrates that BsGH7-3 is an endoglucanase with a long half-life and no loss in activity in the presence of denaturants like salt and ionic liquids, and lays the foundation towards exploring the Bipolaris genome for other cell wall-degrading enzymes. Electronic supplementary material The online version of this article (doi:10.1186/s13068-017-0822-0) contains supplementary material, which is available to authorized users. |
| Databáze: | OpenAIRE |
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