The functionalization methods of materials based on bacterial cellulose (BC) mainly focus on the chemical modification or physical coating of fermentation products, which may cause several problems, such as environment pollution, low reaction efficiency and easy loss of functional moieties during application. Here, we develop a modification method utilizing the in situ microbial fermentation method combined with 6-carboxyfluorescein-modified glucose (6CF-Glc) as a substrate using Komagataeibacter sucrofermentans to produce functional BC with a nonnatural characteristic fluorescence. Our results indicate that the microbial synthesis method is more efficient, controllable and environmentally friendly than traditional modification methods. Therefore, this work confirms that BC can be functionalized by using a microbial synthesis system with functionalized glucose, which provides insights not only for the functionalization of BC but also for the in situ synthesis of other functional materials through microbial synthetic systems.

Natural rubber is a kind of indispensable biopolymers with great use and strategic importance in human society. However, its production relies almost exclusively on rubber-producing plants Hevea brasiliensis, which have high requirements for growth conditions, and the mechanism of natural rubber biosynthesis remains largely unknown. In the past two decades, details of the rubber chain polymerization and proteins involved in natural rubber biosynthesis have been investigated intensively. Meanwhile, omics and other advanced biotechnologies bring new insight into rubber production and development of new rubber-producing plants. This review summarizes the achievements of the past two decades in understanding the biosynthesis of natural rubber, especially the massive information obtained from the omics analyses. Possibilities of natural rubber biosynthesis in vitro or in genetically engineered microorganisms are also discussed.

Background: α-Pinene is an important natural product that is widely used in flavorings, fragrances, medicines, fine chemicals and high-density renewable fuels. Currently, α-Pinene used in industry is mainly produced either by tapping trees (gum turpentine) or as a byproduct of paper pulping (crude sulfate turpentine, CST). However, the extraction of it from trees is tedious and inefficient and requires substantial expenditure of natural resources. Therefore, it is necessary to seek sustainable technologies for α-pinene production. Results: To construct the microbial synthetic pathway of α-pinene in E. coli, we co-expressed native geranyl diphosphate synthase (IspA) from E. coli and α-pinene synthase (Pt30) from Pinus taeda, and then to increase the geranyl diphosphate (GPP) content in the cells, a suitable geranyl diphosphate synthase (GPPS2) was selected from two different origins. Furthermore, to enhance α-pinene production, a novel biosynthetic pathway of α-pinene was assembled in E. coli BL21(DE3) with the heterologous hybrid mevalonate (MVA) pathway, GPPS2 and α-pinene synthase (Pt30). The final genetic strain, YJM28, harboring the above novel biosynthetic pathway of α-pinene, accumulated α-pinene up to 5.44 mg/L and 0.97 g/L under flask and fed-batch fermentation conditions, respectively. The conversion efficiency of glucose to α-pinene (gram to gram) in the metabolically engineered strain reached 2.61%. Conclusions: In this paper, by using metabolic engineering techniques, the more efficient biosynthetic pathway of α-pinene was successfully assembled in E. coli BL21(DE3) with the heterologous hybrid MVA pathway, GPPS2 and α-pinene synthase (Pt30). In addition, this is the first report on α-pinene fed-batch fermentation, and our results represent improvements over previous reports.

This review covers the strategies mostly developed in the last three years for microbial production of isoprenoid, classified according to the engineering targets.

Background: The isopentenols, including isoprenol and prenol, are excellent alternative fuels. However, they are not compounds largely accumulated in natural organism. The need for the next generation of biofuels with better physical and chemical properties impels us to develop biosynthetic routes for the production of isoprenol and prenol from renewable sugar. In this study, we use the heterogenous mevalonate-dependent (MVA) isoprenoid pathway for the synthesis of isopentenyl pyrophosphate (IPP) and dimethylallyl pyrophosphate (DMAPP) intermediates, and then convert IPP and DMAPP to isoprenol and prenol, respectively. Results: A mevalonate titer of 1.7 g/L was obtained by constructing an efficient MVA upper pathway in engineered E. coli. Different phosphatases and pyrophosphatases were investigated for their abilities in hydrolyzing the IPP and DMAPP. Consequently, ADP-ribose pyrophosphatase was found to be an efficient IPP and DMAPP hydrolase. Moreover, ADP-ribose pyrophosphatase from Bacillus subtilis (BsNudF) exhibited a equivalent substrate specificity towards IPP and DMAPP, while ADP-ribose pyrophosphatase from E. coli (EcNudF) presented a high substrate preference for DMAPP. Without the expression of any phosphatases or pyrophosphatases, a background level of isopentenols was synthesized. When the endogenous pyrophosphatase genes (EcNudF and yggV) that were capable of enhancing the hydrolyzation of the IPP and DMAPP were knocked out, the background level of isopentenols was still obtained. Maybe the synthesized IPP and DMAPP were hydrolyzed by some unknown hydrolases of E. coli. Finally, 1.3 g/L single isoprenol was obtained by blocking the conversion of IPP to DMAPP and employing the BsNudF, and 0.2 g/L ~80% prenol was produced by employing the EcNudF. A maximal yield of 12% was achieved in both isoprenol and prenol producing strains. Conclusions: To the best of our knowledge, this is the first successful report on high-specificity production of isoprenol and prenol by microbial fermentation. Over 1.3 g/L isoprenol achieved in shake-flask experiments represents a quite encouraging titer of higher alcohols. In addition, the substrate specificities of ADP-ribose pyrophosphatases were determined and successfully applied for the high-specificity synthesis of isoprenol and prenol. Altogether, this work presents a promising strategy for high-specificity production of two excellent biofuels, isoprenol and prenol.