New lignocellulose integrated biological saccharification process

The economic and cleanliness of lignocellulose biotransformation has always been the biggest bottleneck hindering the large-scale utilization of agricultural and forestry waste such as straw. At present, the mainstream strategy of lignocellulosic biotransformation is the simultaneous saccharification and fermentation process based on free cellulase, but the core enzyme technology is monopolized by foreign companies, and it is difficult to further reduce the cost of enzymes, making the current process not market competitive. Consolidated bioprocessing (CBP) is a lignocellulose conversion strategy proposed in recent years. It integrates cellulase production, lignocellulosic substrate enzymolysis, and final product fermentation into the same reactor. Streamline processes, reduce costs and equipment requirements. However, because the CBP strategy performs multiple steps in the same reactor at the same time, it is necessary to compromise the reaction conditions, it is difficult to obtain high levels of enzyme production, enzymolysis and fermentation at the same time, and the final product is single and difficult to adjust, which greatly limits the Its scope of application.

To this end, the Metabolomics Research Group of the Qingdao Institute of Bioenergy and Process of the Chinese Academy of Sciences has taken a different approach and proposed a new strategy for Consolidated Bio-Saccharification (CBS) based on the whole-cell fiber catalyst. et al, 2019, 12 (1): 35]. This strategy organically integrates the enzyme production and hydrolysis steps, and at the same time separates the downstream fermentation step to a certain degree. The platform compound, which is a fermentable sugar, is used as the target product. It has the advantages of low enzyme cost, simple process, and flexible downstream export. , Is expected to be widely used in lignocellulose biotransformation to produce various chemicals, functional foods and pharmaceuticals.

Fibrosomes are a supramolecular complex secreted by anaerobic microorganisms, such as Clostridium thermocellum, that can efficiently degrade lignocellulose, and are one of the most efficient lignocellulose degradation systems known in nature. The metabolomics research group has long been committed to the genetic modification, mechanism of action, and metabolic engineering research of cellulose-degrading bacteria such as Clostridium thermocellum and their fibrosomes. The genetic manipulation hardware equipment and software tools independently developed by the research group are The physiology and biochemistry of C. cellulosae, the assembly of fibrosomes, synthesis regulation and product inhibition, product uptake and metabolism have been systematically studied, and a deep understanding of the functional mechanism and regulation of C. thermocellum and its fibrosomes has been obtained. . Based on this, the research group first successfully developed a whole-cell catalyst based on fibrosomes in 2017 to achieve the efficient conversion of lignocellulosic substrates to fermentable sugars [Zhang J, et al, 2017,10 (1) : 124], the rudiment of the CBS process has been initially established, but there are still problems such as failure to fully remove product inhibition, decline in key enzyme production, and slow saccharification.

In response to these problems, the research team optimized the fibrosomes by re-in situ reconstruction, and constructed a second-generation whole-bacterial biocatalyst, which affects the process efficiency, medium composition, inoculation volume, seed culture, and substrate load. The key factors are optimized, which significantly improves the saccharification efficiency and shortens the saccharification time. Under the optimal conditions, using the pretreated wheat straw as a substrate, the entire saccharification process time was shortened by 50%, the sugar yield reached 0.795 g / g, and the sugar yield was 89.3% (see figure). This process combined with the matching lignocellulose pretreatment process developed by this laboratory has greatly reduced the cost of lignocellulose saccharification, and has already had the possibility of industrial application.

This work provided a new whole bacteria biocatalyst and an optimized process matched with it for the realization of CBS, which proved that CBS is a feasible low-cost and efficient utilization strategy of lignocellulose. The metabolomics research group reported the process strategy in a research paper published online on February 18 in Biotechnology for Biofuels and formally proposed the concept of "Integrated Biosaccharification (CBS)". The metabolomics research group is currently conducting pilot scale-up of the CBS process and establishing an industrial demonstration system based on this process, which is expected to greatly promote the industrialization of lignocellulose biotransformation.

The doctoral student Liu Shiyue and associate researcher Liu Yajun of this research group co-authored the paper. Researcher Cui Qiu is the corresponding author of the paper. This work was funded by the Strategic Pilot Project of the Chinese Academy of Sciences, the National Natural Science Foundation of China, and the Natural Science Foundation of Shandong Province.