Rethinking Biosynthesis of Aclacinomycin A.

Aclacinomycin A (ACM-A) is an anthracycline antitumor agent widely used in clinical practice. The current industrial production of ACM-A relies primarily on chemical synthesis and microbial fermentation. However, chemical synthesis involves multiple reactions which give rise to high production costs and environmental pollution.

CRISPR-Cas13 technology portfolio and alliance with other genetic tools.

The exploitation of CRISPR-Cas systems especially CRISPR-Cas9 has led to radical advances in genome editing, gene activation, gene repression, protein imaging, and beyond. However, these applications are limited to targeting of DNA rather than RNA. CRISPR-Cas13 is the first reported CRISPR system targeting RNA and thus opens a new avenue for transcription regulation.

Genomic landscapes of bacterial transposons and their applications in strain improvement.

Transposons are mobile genetic elements that can give rise to gene mutation and genome rearrangement. Due to their mobility, transposons have been exploited as genetic tools for modification of plants, animals, and microbes. Although a plethora of reviews have summarized families of transposons, the transposons from fermentation bacteria have not been systematically documented, which thereby constrain the exploitation for metabolic engineering and synthetic biology purposes.

Delineating biosynthesis of Huperzine A, A plant-derived medicine for the treatment of Alzheimer's disease.

Huperzine A (HupA) is a plant-derived lycopodium alkaloid used for the treatment of Alzheimer's disease due to its inhibition against acetylcholinesterase. Currently, industrial production of HupA relies primarily on direct extraction from Huperzia serrate, a perennial herbaceous plant. However, this strategy cannot satisfy the increasing demand for HupA due to scarcity of H.

Engineering Glucose-to-Glycerol Pathway in and Boosting 3-Hydroxypropionic Acid Production Through CRISPR Interference.

The recent decline of the international biodiesel industry has led to decreased production and therefore increased the price of glycerol, which is a major by-product of biodiesel but a substrate for production of 3-hydroxypropionic acid (3-HP), that is, glycerol as a feedstock has no advantage over glucose in price. Hence, we engineered glucose to the glycerol pathway and improved 3-HP production by CRISPR interference (CRISPRi). To begin with, we cloned the genes encoding glycerol 3-phosphate dehydrogenase () and glycerol 3-phosphatase () from , which jointly catalyze glucose into glycerol.

Charting the landscape of RNA polymerases to unleash their potential in strain improvement.

One major mission of microbial cell factory is overproduction of desired chemicals. To this end, it is necessary to orchestrate enzymes that affect metabolic fluxes. However, only modification of a small number of enzymes in most cases cannot maximize desired metabolites, and global regulation is required.

Biosynthesis pathways and strategies for improving 3-hydroxypropionic acid production in bacteria.

3-Hydroxypropionic acid (3-HP) represents an economically important platform compound from which a panel of bulk chemicals can be derived. Compared with petroleum-dependent chemical synthesis, bioproduction of 3-HP has attracted more attention due to utilization of renewable biomass. This review outlines bacterial production of 3-HP, covering aspects of host strains (e.

[Pathway design and key enzyme analysis of diosgenin biosynthesis].

As a naturally occurring steroid sapogenin, diosgenin acts as the precursor of hundreds of steroid medicines, and thereby has important medicinal value. Currently, industrial production of diosgenin relies primarily on chemical extraction from plant materials. Clearly, this strategy shows drawbacks of excessive reliance on plant materials and farmland as well as environment pollution.

Switching metabolic flux by engineering tryptophan operon-assisted CRISPR interference system in Klebsiella pneumoniae.

One grand challenge for bioproduction of desired metabolites is how to coordinate cell growth and product synthesis. Here we report that a tryptophan operon-assisted CRISPR interference (CRISPRi) system can switch glycerol oxidation and reduction pathways in Klebsiella pneumoniae, whereby the oxidation pathway provides energy to sustain growth, and the reduction pathway generates 1,3-propanediol and 3-hydroxypropionic acid (3-HP), two economically important chemicals. Reverse transcription and quantitative PCR (RT-qPCR) showed that this CRISPRi-dependent switch affected the expression of glycerol metabolism-related genes and in turn improved 3-HP production.

A rapid liquid chromatography-electrospray ionization-ion mobility spectrometry method for monitoring nine representative metabolites in the seedlings of cucumber and wheat.

A fast and convenient high-performance liquid chromatography-electrospray ionization-ion mobility spectrometry method was developed to determine nine representative metabolites in the seedlings of cucumber and wheat. The analytical conditions were obtained by optimizing the parameters of high-performance liquid chromatography and ion mobility spectrometry. Briefly, acetonitrile-0.

Intensifying niacin-based biosynthesis of NAD to enhance 3-hydroxypropionic acid production in Klebsiella pneumoniae.

Objective: Glycerol-based biosynthesis of 3-hydroxypropionic acid (3-HP) in Klebsiella pneumoniae involves two reactions: glycerol conversion to 3-hydroxypropionaldehyde (3-HPA) by glycerol dehydratase, and 3-HPA conversion to 3-HP by aldehyde dehydrogenase (ALDH). The ALDH catalysis consumes a lot of cofactor nicotinamide adenine dinucleotide (NAD), which constrains 3-HP production.

Results: Here we report that intensifying niacin-based biosynthesis of NAD can substantially enhance 3-HP production.

CRISPR interference-guided modulation of glucose pathways to boost aconitic acid production in Escherichia coli.

Background: One major mission of microbial breeding is high-level production of desired metabolites. Overproduction of intermediate metabolites in core pathways is challenging as it may impair cell growth and viability.

Results: Here we report that aconitic acid, an intermediate metabolite in tricarboxylic acid (TCA) cycle, can be overproduced by an engineered CRISPR interference (CRISPRi) system in Escherichia coli.

Overview of Membrane Proteins.

species are mainly saprophytic fungi, but some pathotypes of infect economically important plants including cereal crops, vegetables and fruits. Specially, . generates toxins which contaminate food and feed.

Development of orthogonal T7 expression system in Klebsiella pneumoniae.

Most expression systems are tailored for model organisms rather than nonmodel organisms. However, heterologous gene expression in model organisms constrains the innate advantages of original strain carrying gene of interest. In this study, T7 expression system was developed in nonmodel bacterium Klebsiella pneumoniae for production of chemicals.

Improved Production of Pyrroloquinoline Quinone by Simultaneous Augmentation of Its Synthesis Gene Expression and Glucose Metabolism in Klebsiella pneumoniae.

Klebsiella pneumoniae can naturally synthesize pyrroloquinoline quinone (PQQ), but current low yield restricts its commercialization. Here, we reported that PQQ production can be improved by simultaneously intensifying PQQ gene expression and glucose metabolism. Firstly, tandem repetitive tac promoters were constructed to overexpress PQQ synthesis genes.

Engineering a CRISPR Interference System To Repress a Class 1 Integron in Escherichia coli.

Microbial multidrug resistance (MDR) poses a huge threat to human health. Bacterial acquisition of MDR relies primarily on class 1 integron-involved horizontal gene transfer (HGT) of antibiotic resistance genes (ARGs). To date, no strategies other than the use of antibiotics can efficiently cope with MDR.

Physiological investigations of the influences of byproduct pathways on 3-hydroxypropionic acid production in Klebsiella pneumoniae.

Klebsiella pneumoniae can naturally synthesize 3-hydroxypropionic acid (3-HP), 1,3-propanediol (1,3-PD), and 2,3-butanediol (2,3-BD) from glycerol. However, biosynthesis of these industrially important chemicals is constrained by troublesome byproducts. To clarify the influences of byproducts on 3-HP production, in this study, a total of eight byproduct-producing enzyme genes including pmd, poxB, frdB, fumC, dhaT, ilvH, adhP, and pflB were individually deleted from the K.

Exploiting tandem repetitive promoters for high-level production of 3-hydroxypropionic acid.

High-level biosynthesis of desired metabolites is challenging due to complexity of metabolic networks. Here, we report that platform chemical 3-hydroxypropionic acid (3-HP) can be overproduced through promoter engineering and growth-sustaining cultivation, two parallel strategies relying on RNA polymerases (RNAPs). First, we screened a promoter library and revealed that IPTG-inducible tac promoter was most effective for overexpression of PuuC, an endogenous aldehyde dehydrogenase catalyzing 3-HP biosynthesis in Klebsiella pneumoniae.

Engineering CRISPR interference system in Klebsiella pneumoniae for attenuating lactic acid synthesis.

Background: Klebsiella pneumoniae is a promising industrial species for bioproduction of bulk chemicals such as 1,3-propanediol, 2,3-butanediol and 3-hydroxypropionic acid (3-HP). However, lactic acid is a troublesome by-product when optimizing for 3-HP production. Therefore, it is highly desirable to minimize lactic acid.

Development of cyclic AMP receptor protein-based artificial transcription factor for intensifying gene expression.

Vector-dependent gene overexpression typically relies on an efficient operon and sufficient RNA polymerases (RNAPs). The lac (lactose) operon is a paradigm of transcription control, and cyclic AMP receptor protein (CRP) is a global regulator capable of recruiting RNAPs. However, the gap between lac operon and CRP has not been well bridged.

Fundamental CRISPR-Cas9 tools and current applications in microbial systems.

Derived from the bacterial adaptive immune system, CRISPR technology has revolutionized conventional genetic engineering methods and unprecedentedly facilitated strain engineering. In this review, we outline the fundamental CRISPR tools that have been employed for strain optimization. These tools include CRISPR editing, CRISPR interference, CRISPR activation and protein imaging.

Engineering Plasmid-Free Klebsiella Pneumoniae for Production of 3-Hydroxypropionic Acid.

Although plasmid-dependent microbial breeding is predominant in manufacturing bio-based chemicals, it shows pitfall of genetic instability and thus hinders industrial production. Alternatively, chromosome engineering free from plasmid enables genetic stability and thus represents the new trend of microbial breeding. 3-Hydroxypropionic acid (3-HP) is an economically important platform compound as the versatile precursor of a suite of C3 compound, such as 1, 3-PDO.

Enhanced Promoter Activity by Replenishment of Sigma Factor rpoE in Klebsiella pneumoniae.

Plasmid-dependent overexpression of enzyme(s) aims to divert carbon flux toward a desired compound. One drawback of this strategy is compromise of growth due to massive consumption of host resources. Here we show that replenishment of sigma factor rpoE improves the growth of Klebsiella pneumoniae.

High Production of 3-Hydroxypropionic Acid in Klebsiella pneumoniae by Systematic Optimization of Glycerol Metabolism.

3-Hydroxypropionic acid (3-HP) is an important platform chemical proposed by the United States Department of Energy. 3-HP can be converted to a series of bulk chemicals. Biological production of 3-HP has made great progress in recent years.

[Research advances in the virulence factors of Klebsiella pneumoniae--A review].

Klebsiella pneumoniae is of great attractiveness because it naturally produces a series of bulk chemicals such as 1,3-propanediol, 2,3-butanediol and 3-hydroxypropionic acid. Although this species has been fueled in recent years, its pathogenicity is considered an obstacle hindering industrial applications. Here we portray a picture of the virulence factors, including pili, receptors, capsular polysaccharides, lipopolysaccharides, and newly identified virulence factors.