Expression of furfural reductase improved furfural tolerance in Antarctic bacterium pseudomonas extremaustralis

Whole-cell biocatalysis using Antarctic bacteria is presently hampered by a lack of genetic information, limited gene tools and critically, a poor range of cultivation conditions. In this work, biological engineering strategy was employed for developing Pseudomonas extremaustralis, a metabolically-versatile and biopolymer-producing Antarctic bacterium, as a new whole-cell biocatalytic host. For this purpose, gene cloning and plasmid construction were carried out for overexpression of furfural reductase (FucO), an industrially-important enzyme for degradation of toxic furfural compound commonly found in lignocellulosic biorefinery. FucO gene from Escherichia coli BL21 was cloned in pJM105 plasmid and transformed into competent cells of P. extremaustralis to generate a biologically-engineered pFucO strain. For functional characterization of the enzyme, furfural reductase activity was assayed, where the P. extremaustralis pFucO strain exhibited increased furfural reductase activity of about 15.6 U/mg, an 18.8-fold higher than empty plasmid-carrying control pJM105 strain (0.83 U/mg). Furfural detoxification activity using whole cells was also determined by which the overexpression of FucO led to increased tolerance and cell growth with an OD600 value of 5.3 as compared to the control pJM105 strain that was inhibited with 10 mM furfural during 48-hour cultivation. Therefore, the findings obtained in this study successfully demonstrated the development of P. extremaustralis as biocatalytic host for the production of recombinant furfural reductase. The bioengineering would serve as a modular biotechnological platform for polar strain and bioproduct development tailored towards industrial biotechnology applications

Unveiling the Core Effector Proteins of Oil Palm Pathogen Ganoderma boninense via Pan-Secretome Analysis

Ganoderma boninense is the major causal agent of basal stem rot (BSR) disease in oil palm, causing the progressive rot of the basal part of the stem. Despite its prominence, the key pathogenicity determinants for the aggressive nature of hemibiotrophic infection remain unknown. In this study, genome sequencing and the annotation of G. boninense T10 were carried out using the Illumina sequencing platform, and comparative genome analysis was performed with previously reported G. boninense strains (NJ3 and G3). The pan-secretome of G. boninense was constructed and comprised 937 core orthogroups, 243 accessory orthogroups, and 84 strain-specific orthogroups. In total, 320 core orthogroups were enriched with candidate effector proteins (CEPs) that could be classified as carbohydrate-active enzymes, hydrolases, and non-catalytic proteins. Differential expression analysis revealed an upregulation of five CEP genes that was linked to the suppression of PTI signaling cascade, while the downregulation of four CEP genes was linked to the inhibition of PTI by preventing host defense elicitation. Genome architecture analysis revealed the one-speed architecture of the G. boninense genome and the lack of preferential association of CEP genes to transposable elements. The findings obtained from this study aid in the characterization of pathogenicity determinants and molecular biomarkers of BSR disease