Genomic Features and Construction of Streamlined Genome Chassis of Nisin Z Producer Lactococcus lactis N8

Lactococcus lactis is a commonly used fermenting bacteria in cheese, beverages and meat products. Due to the lack of simplified chassis strains, it has not been widely used in the fields of synthetic biology. Thus, the construction of lactic acid bacteria chassis strains becomes more and more important. In this study, we performed whole genome sequencing, annotation and analysis of L. lactis N8. Based on the genome analysis, we found that L. lactis N8 contains two large plasmids, and the function prediction of the plasmids shows that some regions are related to carbohydrate transport/metabolism, multi-stress resistance and amino acid uptake. L. lactis N8 contains a total of seven prophage-related fragments and twelve genomic islands. A gene cluster encoding a hybrid NRPS–PKS system that was found in L. lactis N8 reveals that the strain has the potential to synthesize novel secondary metabolites. Furthermore, we have constructed a simplified genome chassis of L. lactis N8 and achieved the largest amount of deletion of L. lactis so far. Taken together, the present study offers further insights into the function and potential role of L. lactis N8 as a model strain of lactic acid bacteria and lays the foundation for its application in the field of synthetic biology

Genomic Features and Construction of Streamlined Genome Chassis of Nisin Z Producer Lactococcus lactis N8

Lactococcus lactis is a commonly used fermenting bacteria in cheese, beverages and meat products. Due to the lack of simplified chassis strains, it has not been widely used in the fields of synthetic biology. Thus, the construction of lactic acid bacteria chassis strains becomes more and more important. In this study, we performed whole genome sequencing, annotation and analysis of L. lactis N8. Based on the genome analysis, we found that L. lactis N8 contains two large plasmids, and the function prediction of the plasmids shows that some regions are related to carbohydrate transport/metabolism, multi-stress resistance and amino acid uptake. L. lactis N8 contains a total of seven prophage-related fragments and twelve genomic islands. A gene cluster encoding a hybrid NRPS–PKS system that was found in L. lactis N8 reveals that the strain has the potential to synthesize novel secondary metabolites. Furthermore, we have constructed a simplified genome chassis of L. lactis N8 and achieved the largest amount of deletion of L. lactis so far. Taken together, the present study offers further insights into the function and potential role of L. lactis N8 as a model strain of lactic acid bacteria and lays the foundation for its application in the field of synthetic biology

Engineering Lactococcus lactis as a multi-stress tolerant biosynthetic chassis by deleting the prophage-related fragment

Background In bioengineering, growth of microorganisms is limited because of environmental and industrial stresses during fermentation. This study aimed to construct a nisin-producing chassis Lactococcus lactis strain with genome-streamlined, low metabolic burden, and multi-stress tolerance characteristics. Results The Cre-loxP recombination system was applied to reduce the genome and obtain the target chassis strain. A prophage-related fragment (PRF; 19,739 bp) in the L. lactis N8 genome was deleted, and the mutant strain L. lactis N8-1 was chosen for multi-stress tolerance studies. Nisin immunity of L. lactis N8-1 was increased to 6500 IU/mL, which was 44.44% higher than that of the wild-type L. lactis N8 (4500 IU/mL). The survival rates of L. lactis N8-1 treated with lysozyme for 2 h and lactic acid for 1 h were 1000- and 10,000-fold higher than that of the wild-type strain, respectively. At 39 celcius, the L. lactis N8-1 could still maintain its growth, whereas the growth of the wild-type strain dramatically dropped. Scanning electron microscopy showed that the cell wall integrity of L. lactis N8-1 was well maintained after lysozyme treatment. Tandem mass tags labeled quantitative proteomics revealed that 33 and 9 proteins were significantly upregulated and downregulated, respectively, in L. lactis N8-1. These differential proteins were involved in carbohydrate and energy transport/metabolism, biosynthesis of cell wall and cell surface proteins. Conclusions PRF deletion was proven to be an efficient strategy to achieve multi-stress tolerance and nisin immunity in L. lactis, thereby providing a new perspective for industrially obtaining engineered strains with multi-stress tolerance and expanding the application of lactic acid bacteria in biotechnology and synthetic biology. Besides, the importance of PRF, which can confer vital phenotypes to bacteria, was established.Peer reviewe

Expression and Characterization of a New PolyG-Specific Alginate Lyase From Marine Bacterium Microbulbifer sp. Q7

Alginate lyases play an important role in preparation of alginate oligosaccharides. Although a large number of alginate lyases have been characterized, reports on directional preparation of alginate oligosaccharides by alginate lyases are still rather less. Here, a gene alyM encoding a new alginate lyase AlyM was cloned from Microbulbifer sp. Q7 and expressed in Escherichia coli. AlyM exhibited the maximumactivity at pH 7.0 and 55°C and showed special preference to poly-guluronic acid (polyG). Glycine promoted the extracellular secretion of AlyM by 3.6 times. PBS and glycerol significantly improved the thermal stability of AlyM, the enzyme activity remained 75 and 78% after heat-treatment at 45°C for 2 h, respectively. ESI-MS analysis suggested that AlyM mainly produced oligosaccharides with degrees of polymerization (DP) of 2–5. The results of 1H-NMR showed that guluronic acid (G) occupied the reducing end of the end products, indicating that AlyM preferred to degrade the glycosidic bond at the G-X linkage. HPLC analysis showed that the hydrolysis products with a lower degree of polymerization contained more G. Therefore, AlyM shows good potential to produce alginate oligosaccharides with specific M/G ratio and molecular weights

Unlocking emerging impacts of carbon tax on integrated energy systems through supply and demand co-optimization

Integrated energy systems (IES) can help achieve greater energy efficiency, and then ultimately promote a climate-neutral economy by utilizing local renewable resources. Demand-side energy-saving measures can reduce operational costs associated with energy usage. Most existing IES models, however, focus on supply-side optimization, while the demand-side energy-saving potential and its impacts on whole-system performance are still not clear. The increasing carbon tax makes it even more important to understand the interactions between supply and demand sides to achieve a sustainable system with a minimal carbon charge. Hence, this study proposes a co-optimization model to simultaneously optimize the supply and demand sides of an IES considering the impact of the carbon tax. A selection tree is developed to describe various demand-side envelope upgrading technologies, and a binary tree is established by generating a set of supply-side scenarios with corresponding probabilities. Based on these results, an improved two-stage stochastic programming model is proposed. The robustness of the modeling results was further validated by a simulation–optimization-based uncertainty analysis addressing price uncertainties. A case study in Shanghai indicates that the proposed co-optimization model achieves more cost-efficient solutions than supply-side-only optimization considering carbon tax. Introducing carbon tax can reduce the installed capacity of fuel-based energy technologies by up to 24% and greatly accelerate the penetration of renewables. The increasing carbon tax also promotes the adoption of more advanced energy-saving technologies. Uncertainty analysis reveals acceptable robustness of the optimal demand-side scheme and supply-side configuration with a deviation of less than 5% and a coefficient of variation of 7%. Overall, the observations of the proposed model and case study provide valuable insights for IES design considering an emerging charge of carbon tax

A cartilage-inspired lubrication system

Articular cartilage is an example of a highly efficacious water-based, natural lubrication system that is optimized to provide low friction and wear protection at both low and high loads and sliding velocities. One of the secrets of cartilage\u27s superior tribology comes from a unique, multimodal lubrication strategy consisting of both a fluid pressurization mediated lubrication mechanism and a boundary lubrication mechanism supported by surface bound macromolecules. Using a reconstituted network of highly interconnected cellulose fibers and simple modification through the immobilization of polyelectrolytes, we have recreated many of the mechanical and chemical properties of cartilage and the cartilage lubrication system to produce a purely synthetic material system that exhibits some of the same lubrication mechanisms, time dependent friction response, and high wear resistance as natural cartilage tissue. Friction and wear studies demonstrate how the properties of the cellulose fiber network can be used to control and optimize the lubrication and wear resistance of the material surfaces and highlight what key features of cartilage should be duplicated in order to produce a cartilage-mimetic lubrication system

Community-based study on CKD subjects and the associated risk factors

Background. The study was performed to investigate the prevalence, awareness and the risk factors of chronic kidney disease (CKD) in the community population in Shanghai, China

Assimilation of Typhoon Wind Field Retrieved from Scatterometer and SAR Based on the Huber Norm Quality Control

Observations of sea surface wind field are critical for typhoon prediction. The scatterometer observation is one of the most important sources of sea surface winds, which provides both wind speed and wind direction information. However, the spatial resolution of scatterometer wind is low. Synthetic Aperture Radar (SAR) can provide a more detailed wind structure of the tropical cyclone. In addition, the cross-polarization observation of SAR can provide more detailed information of high speed wind (>25 m·s − 1 ) than the scatterometer. Nevertheless, due to the narrow swath of SAR, the number of retrieved sea surface wind data used in the data assimilation is limited, and another limitation of SAR wind observation is that it does not provide true wind direction information. In this paper, the joint assimilation of the Advanced Scatterometer (ASCAT) wind and Sentinel-1 SAR wind was investigated. Another limitation in the current operational typhoon prediction is the inefficient quality control (QC) method used in the data assimilation since a large number of high speed wind observations was rejected by the traditional Gaussian distribution QC. We introduce the Huber norm distribution quality control (QC) into the data assimilation successfully. A numerical simulation experiment of typhoon by Lionrock (2016) is conducted to test the proposed method. The experimental results showed that the new quality control scheme not only greatly increases the availability of wind data in the area of the typhoon center, but also improves the typhoon track prediction, as well as the intensity prediction. The joint assimilation of scatterometer and SAR winds does have a positive impact on the typhoon prediction