Mevalonate pathway analysis of Saccharomyces cerevisiae during bioisoprene synthesis

Isoprene, synthesized through two complementary biosynthetic routes known as the mevalonate (MVA) pathway and the deoxy-xylulose phosphate pathway, is a valuable monomer that is used for rubber and several other chemical industries. Despite the recent interest in the industrial and biomedical applications of isoprene and its derivatives, the complexity of controlling its chemical synthesis due to the formation of greenhouse gases is a significant problem. To overcome the productivity and yield challenges, in addition to generating environmental and economic benefits, this study aimed to focus on the direct fermentation of cellulosic materials into bioisoprene. In this study, bioisoprene was synthesized via a biotransformation process through enzymatic hydrolysis of cassava peel using Aspergillus niger 11JK and Saccharomyces cerevisiae 19KB strain. The mevalonate (MVA) pathway (synthetic route) exploited during bioisoprene production by S. cerevisiae 19KB strain was investigated using the hydrolyzed cassava peel broth. The obtained crude extract was analyzed for bioisoprene yield and enzymatic activities using Gas chromatography. Furthermore, results of the size exclusion chromatography revealed the presence of polysaccharide hydrolyzing enzymes (e.g., amylase and cellulase), and mevalonate pathway enzymes, including isoprene synthase, mevalonate-5-diphosphate decarboxylase, and isopentyl phosphate kinase, in addition to isoprene, mevalonic acid (MVA), and its isomer dimethylallyl diphosphate (DMAPP). Based on the results obtained in this study, bioisoprene synthesis via direct fermentation of cheap and abundant carbon sources such as cassava peel using the S. cerevisiae 19KB strain will overcome the high production costs and low yield challenges of bioisoprene, thus generating significant environmental and economic benefits

Metabolic potential of Azotobacter alginate producers and sustainable alternatives for alginate extraction

This research aimed to assess the metabolic activities of Azotobacter vinelandii DT1 strain by assaying its alginate synthesizing enzymes and modifying genes, synthesize alginate from the agricultural residues, and eradicate the pathogenicity and risk associated with Pseudomonas aeruginosa alginate. Preliminary screening for alginate production revealed the presence of different alginate products. High-performance liquid chromatography (HPLC) confirmed the existence of varying alginate concentrations, such as sodium alginate (2.42754×10-1 g/ 100 ml), calcium alginate (1.09597×10-1 g/ 100 ml), acid alginate (1.39420×10-2 g/ 100 ml), alginate oligosaccharide (8.20576×10-2 g/100 ml), and potassium alginate (9.78836×10-2 g/ 100 ml). These were accompanied with the corresponding alginate synthesizing enzymes; mainly GDP-Mannose dehydrogenase (23.77± 0.13 U/ ml); glycosyltransferase (9.68± 0.53 U/ ml), phosphomannomutase (266.09± 0.16 U/ ml), mannose phosphate isomerase (95.87± 0.51 U/ ml), alginate lyase (24.50± 0.95 U/ ml), and mannuronan epimerase (49.93± 0.82 U/ ml). In this study, the expression of alginate-modifying genes such as alginate lyase and GDP-Mannose dehydrogenase amplicons of Azotobacter vinelandii DT1 strain corresponding to 766 bp, 43 ng and 600 bp, 33 ng molecular weights of the fast DNA marker; justified the synthesis of different alginate products. Azotobacter alginate synthesis using a low-cost substrate (i.e., corn cobs) and completely non-pathogenic bacteria (Azotobacter vinelandii DT1) may be desirable compared to the pathogenicity risks and poor jellying qualities linked to Pseudomonas alginate biosynthesis, its expensive production costs, and the adverse environmental effects of seaweed harvesting and processing