Microorganisms are ubiquitous in people's daily life, as far as diet is concerned, foods such as bread, monosodium glutamate, soy sauce, beer and other foods are actually the products of microbial fermentation, and behind this, they are inseparable from microbial strains with excellent characteristics.
Strains are the core lifeblood of the microbial industry, just as chips are to the semiconductor industry. In recent years, with the development of genetic engineering, metabolic engineering, gene editing, droplet microfluidics and synthetic biology, new opportunities have been brought for the screening and efficiency improvement of excellent microbial strains.
Recently, Shenghui interviewed Dr. Ye Jianwen, associate professor of the School of Biological Science and Engineering of South China University of Technology, who interpreted the development status and application prospects of microbial breeding, synthetic biology, and manufacturing of new polyhydroxyalkanoate esters (PHAs).
Photo: Dr. Ye Jianwen, Associate Professor, School of Biological Science and Engineering, South China University of Technology (Source: Interviewee)
After graduating from South China University of Technology with a bachelor's degree, Ye Jianwen was sent to Tsinghua University to study for a doctorate (direct doctorate) under the supervision of Professor Chen Guoqiang, and his research topic was the genetic modification and downstream industrial application of microorganisms such as Salinomonas, Pseudomonas, Escherichia coli, Roche, etc., and continued to complete postdoctoral research in Professor Chen Guoqiang's research group.
Today, Ye Jianwen is an associate professor and doctoral supervisor at the School of Biological Science and Engineering at South China University of Technology, and his research interests mainly focus on microbial genetic modification and downstream industrial applications, and continue to expand the "next generation of industrial biotechnology" based on Minemonas (proposed by Professor Chen Guoqiang of Tsinghua University). Up to now, Ye Jianwen has published more than 20 papers in journals such as Nature Communications, Advanced Materials, Trends in Biotechnology, Metabolic Engineering, Bioresource Technology, and Essays in Biochemistry as a corresponding author, and has applied for/authorized 19 invention patents. Lun Shiyi Education Foundation" Young Scholar, Lindau Nobel Prize in Young Science (Chemistry) and other honors.
"To sum up, I have two core keywords throughout my academic career, research and entrepreneurship." Ye Jianwen introduced, "The scientific research mainly focuses on genetic modification of halomonas; In terms of entrepreneurship, it is to put the microorganisms that have been previously studied into factories and carry out large-scale industrial production. Ye Jianwen said frankly that during the entrepreneurial period, he worked sleepless for more than 70 hours in the pilot plant, "Now looking back, I cherish and appreciate that time very much, Professor Chen and others have given me a broad space and platform, and I have been well trained and grown in all aspects." He said.
Microbial breeding is an important support for the biomanufacturing industry
Similar to plant breeding, microbial breeding is a biological technology for cultivating excellent microorganisms, which can be roughly divided into two categories, natural breeding and artificial selection. "Natural breeding is to screen some characteristic microorganisms or chassis bacteria in nature, while artificial breeding is to mutate and breed microorganisms through chemical mutagenesis, plasma mutagenesis and other methods. These two breeding styles can be combined or independent. Ye Jianwen said.
"When genetic modification is difficult to make a breakthrough in the short term or faces a theoretical bottleneck that is difficult to break through, a characteristic chassis based on natural selection can be selected and functionally strengthened. This involves genetic modification, which is centered on studying, intensifying, and refining its functional molecular or metabolic mechanisms, and even 'transplanting' them into a traditional model chassis to achieve the same function. He said.
In terms of prospects, in Ye Jianwen's view, microorganisms are an important chassis choice for biosynthesis, so microbial breeding has broad application prospects in the future biomanufacturing industry, and it is also used as a basic technology to support the development of the biomanufacturing industry.
The application prospect is generally promising, but at this stage, including the design of high-throughput and screening processes, as well as the genetic stability and editability of artificially selected strains, are all challenges faced by microbial breeding. "Although there is no one-size-fits-all technology system that can overcome all the challenges, there is a point where we are making a technological breakthrough, so we can gradually build the system and solve the various challenges we are currently facing through the 'break-through' route." Ye Jianwen said.
As a technical means, one of the important applications of breeding excellent microbial strains is the efficient synthesis and manufacturing of new biomaterials. Now, with the escalation of the plastic restriction order and the proposal of carbon neutrality and carbon peaking goals, the industrial biotechnology of synthesizing polyhydroxyalkanoates (PHAs) from genetically engineered microorganisms has attracted much attention from the market.
PHA is a kind of polymer polyester material synthesized by microorganisms with good biocompatibility and degradability. According to reports, as a new type of biomaterial, the production and manufacturing process of PHA is fully biosynthetic, which is green and sustainable, and its fermentation conditions are relatively mild. At the same time, PHA itself is completely biodegradable, and it is currently a polyester material that can be fully biodegradable under natural conditions among bio-based materials.
For the challenges faced by PHA biomanufacturing and processing at this stage, Ye Jianwen summarized three aspects.
First, the cost, i.e., how to reduce the production cost of PHA to a level roughly equivalent to that of chemical production of plastics;
Secondly, stability, that is, how to ensure the stable and uniform performance of batch discharge in the fermentation process. "PHA is a material synthesized in living systems, and due to the differences in each cell, it is difficult to ensure that the performance of PHA synthesized by all cells is exactly the same, for example, the yield and yield of each cell, and whether the physicochemical and thermodynamic properties of the synthesized PHA can be maintained." Ye Jianwen said.
Thirdly, the industrial supporting (manufacturing and processing) around the PHA. "After all, PHA is a new material that has just entered the market, so it is very important to quickly match the needs of large-scale industries such as fermentation production, downstream processing and modification." He noted.
In addition to this, the production process is also a major challenge today. "Different synthesis technologies represented by different chassis cells such as Salinomonas, Escherichia coli, and Roche have their own characteristics, and the advantages and disadvantages can only be shown after comprehensive consideration after engineering integration. How to integrate process characteristics, use engineering concepts, shift the balance between technology and economy, and realize the global optimization and continuous technology iteration of the process system is the key to determining the future cost, stability and scale success of PHA. He said.
In this regard, Ye Jianwen believes that the key to solving the problem lies in the analysis of production costs and technical systems, and then the entire process system is optimized, and the corresponding strains or process transformation designs are matched at the same time. "At present, the PHA production process is a completely new process route, which needs to be continuously focused from large to small. At the upstream level, strain optimization is imperative, which not only determines the conversion rate, but also determines the production intensity and material stability. At the downstream level, production processes and material processing also need to be matched and coordinated. He said.
The large-scale production of any product needs to go through a scale-up process, and cost is a key point for PHA, so it is necessary to optimize and expand the process with cost reduction as the core, so as to achieve the orderly development of small-scale to large-scale production. "As a new type of green material, and also one of the most recognizable products in the development of synthetic biology, the application scenarios of PHA are very popular, covering a variety of application markets in the low, medium and high-end, and the application prospects of PHA are promising in the future." Ye Jianwen pointed out.
Engineered Synthesis: PHA and high-value products from Anthropomonas (Source: Essays in Biochemistry)
Whether it's microbial breeding or PHA biosynthesis, these are actually in the category of synthetic biology. As an important sub-discipline of multidisciplinary interdisciplinary incubation of biological sciences in this century, synthetic biology has risen rapidly. As far as China is concerned, Ye Jianwen said, "In recent years, the field of synthetic biology in China has also achieved rapid development, among which, 'fast' is more reflected in the upstream technology level. "For example, gene editing, DNA synthesis, DNA sequencing and other technical means." However, compared with the rapid development of upstream technology, the downstream industry matching is relatively late-aware, including industry/engineering matching, production equipment/integrated system matching, etc. In addition, around the biomanufacturing system, the technical economy, engineering feasibility and stability evaluation need to be further strengthened. He added.
In addition to the technical level, another important factor is the cultivation and construction of the talent training system. "It is very important to understand both upstream technology development and downstream engineering production, and the construction of a compound talent training system will be conducive to the implementation of synthetic biology industry." Ye Jianwen pointed out.
"On the whole, the entire field of synthetic biology is thriving, with a lot of room for imagination, and the future is promising." Talking about the future development prospects, Ye Jianwen summarized three "degrees": we should not only examine the progress of synthetic biology technology from a pessimistic perspective, but also do a good job in the current research work from an optimistic attitude, and in addition, we should also think about the relationship between technological progress, individual scientific research workers and social development from the depth of humanities. "With these reflections, we can promote and contribute to the healthy development of technology and market competition, which is also the concept of 'humanistic innovation' advocated by Professor Qiu Yong during my study at Tsinghua University. In fact, science and engineering disciplines need to have humanistic thinking, only in this way can we better assist us to do a good job in technology development, and at the same time, we will not forget the original intention of the entire industrial ecological construction. Ye Jianwen said.
Engineering is an inevitable expansion process for scientific research, and cost and stability are the key
At this stage, Ye Jianwen is leading the team to develop innovative technologies around extreme chassis microorganisms, such as metabolic engineering, biological fermentation and matching downstream process scale-up technology systems, as well as biosynthesis of multi-product pipelines based on new chassis technologies, so as to achieve a more efficient, green and low-carbon biomanufacturing process.
"The green biomanufacturing process involves the production of downstream such as functional PHA and high value-added amino acid derivatives, as well as the upstream comprehensive utilization system of carbon sources such as carbon dioxide and biomass. Among them, the core is gene editing, strain modification, and the downstream biological fermentation, separation and purification process that match, and these two parts are bound to be closely connected. Ye Jianwen pointed out.
Comparison between traditional industrial biotechnology and new generation industrial biotechnology (Source: Chinese Journal of Bioprocess Engineering)
Regarding the new research progress of the research group, Ye Jianwen summarized several levels:
First of all, at the breeding level, he and his team took samples in some extreme environments and carried out natural breeding of microbial chassis based on the self-designed breeding process, and carried out fermentation optimization and metabolic enhancement. Among them, metabolic enhancement includes two aspects: on the one hand, it focuses on the metabolic utilization of substrates by strains, and then strengthens them on the basis of the "talent" of strains; The other part revolves around the metabolites of the strain. "Because some chassis prefer or are suitable for synthesizing a certain type of product, we will further strengthen its metabolic pathway." Ye Jianwen explained.
Second, at the product level, it mainly revolves around PHA. "PHA is a large category, which has undergone several generations of optimization, and more diversified polymerization monomer designs can be carried out in the future, such as the molar ratio of different monomers may affect the performance of PHA materials, and then the control of PHA polymerization process, etc., we are also currently carrying out research work on PHA synthesis." He said. "In addition to PHA, we have also made some new progress in the biosynthesis of amino acid derivatives, and at this stage we are building a comprehensive utilization system with carbon dioxide as a carbon source, and a relatively mature engineering technology system has been formed."
Talking about the next research trend of the research group, Ye Jianwen said that the next plan is to build an integrated technology platform of "production, education and research" based on Minemonas, supplemented by model organisms, and guided by engineering technology application.
According to reports, the reason why model organisms should be supplemented is because the metabolic characteristics, genomic information and gene editing modification of halomonas need to be supported and assisted by the research background and information of model organisms, "so we will also carry out basic research to explore the mechanism of model organisms and the commonality of gene editing, so as to better study halomonas." He said.
At the same time, "while doing a good job in scientific research, we must not lose the engineering part." Ye Jianwen pointed out that, for example, a relatively good chassis cell can be built in the laboratory, but it is likely to be stuck by a small problem when it enters the factory for large-scale production, resulting in the entire project and industrialization process being shelved. Therefore, we will consider many industrialization issues while conducting scientific research, explore what potential risk points may exist in downstream engineering technologies, and then combine risk points to reverse the development and matching of upstream cell factories. He added.
Specific to the implementation of the industry, in Ye Jianwen's view, the core technical issue is stability, such as the stability of the strain itself (such as genetic material, metabolic activities, etc.) and the stability of the strain performance, as well as the stability of the downstream process. "Stability is closely related to the economy and feasibility of large-scale production, and it will also involve the design of downstream processes. He noted.
As for the relationship between laboratory "scientific research" and industrialized "large-scale production", in Ye Jianwen's view, "from scientific research to large-scale production, it will inevitably go through a gradual process of amplification, and this process is like a 'demon mirror' of scientific research." The depth of thinking or technical perfection in the early stage of scientific research will be well reflected in this process. He noted.
Scientific research focuses more on the cutting-edge exploration of technology, mechanism, natural laws, etc., which belongs to the upstream technology development level, and does not need to consider too many factors such as cost and stability. However, in large-scale production, economy and stability are more important, because they are closely related to the market and are directly related to the acceptability of the market. For example, the biosynthesis industry pays more attention to the three gold standards, yield, yield and conversion rate.
"The issues that need to be considered in the process of large-scale production will be more systematic than scientific research, and technological advancement is not the primary factor, more often, some process fine-tuning, equipment iteration and combined innovation are more important in large-scale production." Ye Jianwen said.
When it comes to industrial transformation, Ye Jianwen said frankly that there is still a talent gap in the field of biomanufacturing in China, so at this stage, he mainly returns to his alma mater (South China University of Technology), relying on the engineering platform characteristics of the School of Biology, with scientific research and talent training as the main line, and teaching and research are mutually beneficial. "In the past year and a half of carrying out scientific research in Huagong, we have been focusing on technology research and development and industrial transformation, and have maintained close industry-university-research cooperation with enterprises. At the same time, it is hoped that a preliminary achievement transformation plan can be formed next year, which will produce a positive output for downstream applications and even the entire industry. At the same time, our team is also working with companies to help them achieve greater growth than expected through our newly developed technology, so that the technology can be recognized by the market, generate value, and form synergies that are mutually reinforcing. He concluded.