Protective RBD-dimer vaccines against SARS-CoV-2 and its variants produced in glycoengineered Pichia pastoris.

Protective vaccines are crucial for preventing and controlling coronavirus disease 2019 (COVID-19). Updated vaccines are needed to confront the continuously evolving and circulating severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants. These vaccines should be safe, effective, amenable to easily scalable production, and affordable.

[Progress and perspectives of natural cell factories for chemical production from methanol].

Methanol has been considered one of the most important alternative carbon sources for the next-generation biomanufacturing due to its low price, mature production processes, and potential sustainability. Constructing microbial cell factories for methanol to chemical biotransformation has become a research hotspot in the green biomanufacturing industry. Focusing on the microorganisms that can naturally use methanol, we compare them with non-natural cell factories for chemical production from methanol.

Scramblase activity of proteorhodopsin confers physiological advantages to in the absence of light.

Microbial rhodopsins are widely distributed in the aqua-ecosystem due to their simple structure and multifaceted functions. Conventionally, microbial rhodopsins are considered to be exclusively light active. Here, we report the discovery of light-independent function of a proteorhodopsin from a psychrophile (ptqPR).

Metabolic engineering of methylotrophic Pichia pastoris for the production of β-alanine.

β-Alanine (3-aminopropionic acid) is the only naturally occurring β-amino acid and an important precursor for the synthesis of a variety of nitrogen-containing chemicals. Fermentative production of β-alanine from renewable feedstocks such as glucose has attracted significant interest in recent years. Methanol has become an emerging and promising renewable feedstock for biomanufacturing as an alternative to glucose.

[Heterologous expression and function evaluation of Gloeobacter violaceus rhodopsin in Escherichia coli].

Proton-pumping rhodopsin (PPR) is a simple photosystem widely distributed in nature. By binding to retinal, PPR can transfer protons from the cytoplasmic to the extracellular side of the membrane under illumination, creating a proton motive force (PMF) to synthesize ATP. The conversion of light into chemical energy by introducing rhodopsin into nonphotosynthetic engineered strains could contribute to promoting growth, increasing production and improving cell tolerance of microbial hosts.

High-level production of glucose oxidase in Pichia pastoris: Effects of Hac1p overexpression on cell physiology and enzyme expression.

Secretion is a common bottleneck in the production of industrial proteins. Although overexpression of the unfolded protein response regulator Hac1p has been widely used to enhance protein secretion, its effects on the physiology of host cells were often overlooked, which would attenuate and even neutralize its beneficial effects on overall protein production. In order to achieve high-level glucose oxidase (GOX) production in Pichia pastoris, we used a set of Hac1p homologues from Saccharomyces cerevisiae (ScHac1p), P.

Engineering unnatural methylotrophic cell factories for methanol-based biomanufacturing: Challenges and opportunities.

Methanol is a very promising feedstock alternative to sugar-based raw materials for biomanufacturing because it does not compete with food production, is abundant and potentially sustainable in the future. Although methylotrophic fermentations have been practiced for decades, their applications are limited by technical drawbacks and insufficient knowledge of the physiology and metabolic regulation of native methylotrophs. Synthetic biology offers great opportunities for engineering efficient methylotrophic microbial cell factories by enabling non-methylotrophic model organisms to utilize methanol via the introduction of C1 utilization pathways.

Pichia pastoris as a Versatile Cell Factory for the Production of Industrial Enzymes and Chemicals: Current Status and Future Perspectives.

With nearly three decades of development, Pichia pastoris (P. pastoris) has become a powerful eukaryotic protein expression system for the expression of thousands of proteins both on a laboratory and industrial scale. In addition, it has also been extensively used as a cell factory for the production of a variety of chemicals.

Hac1p homologues from higher eukaryotes can improve the secretion of heterologous proteins in the yeast Pichia pastoris.

Objectives: To systematically explore the effects of overexpressing Hac1p homologues from different sources on protein secretion in Pichia pastoris system.

Results: Effects of Hac1p homologues encompassing P. pastoris (PpHac1p), S.

Viability and Stress Response of Putative Probiotic Lactobacillus plantarum Strains in Honey Environment.

Due to problem of preservation of dairy products which serve as a matrix for probiotics, it is challenging to use these probiotics as food supplements in many developing countries. To determine the suitability of the Lactobacillus strains for exploitation as probiotics in honey, we investigated the effect of their storage on the viability, functionality, and the mechanism associated with their protective effect. Three isolates obtained from our laboratory collection were identified through amplification of the 16S rRNA gene.

Discovery of novel highly active and stable aspartate dehydrogenases.

Aspartate family amino acids (AFAAs) have important commercial values due to their wide spectrum of applications. Most if not all AFAAs are produced under aerobic conditions which is energy-intensive. To establish a cost-effective anaerobic process for production of AFAAs, it holds great promise to develop a new pathway enabling the conversion of oxoloacetate into aspartate through direct amination which is catalyzed by aspartate dehydrogenase (AspDH).

[Optimization of 1,2,4-butanetriol synthetic pathway in Escherichia coli].

1,2,4-Butanetriol (BT) is an important non-natural chemical with a variety of industrial applications. A recombinant Escherichia coli biosynthesizing BT from D-xylose was constructed by heterologously expressing xdh and mdlC, and knocking out competing pathway genes including xylA, xylB, yjhE, yagH and ycdW. To optimize BT synthesis pathway, the third catalytic step that catalyzes the decarboxylation reaction of 3-deoxy-D-glycero-pentulosonic acid was identified as a potential bottleneck.

From cyanochemicals to cyanofactories: a review and perspective.

Engineering cyanobacteria for production of chemicals from solar energy, CO2 and water is a potential approach to address global energy and environment issues such as greenhouse effect. To date, more than 20 chemicals have been synthesized by engineered cyanobacteria using CO2 as raw materials, and these studies have been well reviewed. However, unlike heterotrophic microorganisms, the low CO2 fixation rate makes it a long way to go from cyanochemicals to cyanofactories.

[Recent development of Pichia pastoris system: current status and future perspective].

With more than 20 years of development, Pichia pastoris system has been extensively used both on a lab and industrial scale. This review outlines the progress made on P. pastoris from aspects of protein expression, molecular engineering tools and methods, and biochemical production.

Synthetic pathway optimization for improved 1,2,4-butanetriol production.

1,2,4-Butanetriol (BT) is an important non-natural chemical with a variety of industrial applications. Identifying the bottlenecks for BT biosynthesis is expected to contribute to improving the efficiency of this process. In this work, we first constructed a prototype strain for BT production by assembling a four-step synthetic pathway and disrupting the competing pathways for xylose in Escherichia coli BW25113.

Construction of efficient xylose utilizing Pichia pastoris for industrial enzyme production.

Background: Cellulosic biomass especially agricultural/wood residues can be utilized as feedstock to cost-effectively produce fuels, chemicals and bulk industrial enzymes, which demands xylose utilization from microbial cell factories. While previous works have made significant progress in improving microbial conversion of xylose into fuels and chemicals, no study has reported the engineering of efficient xylose utilizing protein expression systems for the purpose of producing industrial enzymes.

Results: In this work, using Pichia pastoris as an example, we demonstrated the successful engineering of xylose metabolizing ability into of protein expression systems.

[Direct secretory expression of active microbial transglutaminase in Pichia pastoris].

Direct secretory expression of active microbial transglutaminase (MTG) using heterologous hosts is a promising strategy, although its production level still needs to be improved for industrial production. Pichia pastoris is one of the most efficient expression systems developed in recent years. In this study, secretory expression of active MTG was successfully achieved by co-expressing the pro sequence and mature MTG genes in P.

Using Bacillus amyloliquefaciens for remediation of aquaculture water.

Remediation of aquaculture water using microorganisms like Bacillus species is a burgeoning trend for the sustainable development of aquaculture industries. In this work, a Bacillus amyloliquefaciens strain (namely B. amyloliquefaciens HN), isolated from activated sludge of a polluted river, was evaluated for its potential in water remediation using simulated aquaculture water.

Enhancement of the gene targeting efficiency of non-conventional yeasts by increasing genetic redundancy.

In contrast to model yeasts, gene targeting efficiencies of non-conventional yeasts are usually low, which greatly limits the research and applications of these organisms. In this study, we aimed to enhance the gene targeting efficiency of non-conventional yeasts by improving the fitness of mutant strains, particularly by increasing the genetic redundancy of host cells. To demonstrate this process, OCH1 gene deletion in Pichia pastoris was performed.

Transcriptional investigation of the effect of mixed feeding to identify the main cellular stresses on recombinant Pichia pastoris.

Heterologous protein expression using Pichia pastoris causes metabolic stress on the physiology of host cells, which may compromise the yields of secreted foreign proteins. Thus, understanding these metabolic stresses during secretory expression allows us to circumvent these undesirable effects. We investigated the effect of co-feeding two alternative carbon resources, sorbitol and yeast extract (YE), on the physiology of A3, a P.

Expression of bacterial GshF in Pichia pastoris for glutathione production.

Conventionally, two consecutive enzymatic reactions catalyzed by γ-glutamylcysteine synthetase and glutathione synthetase are most commonly used for glutathione production. Here we demonstrate that bacterial bifunctional GshF can be used for glutathione production in a eukaryotic system without accumulation of the intermediate γ-glutamylcysteine.

Combinatorial strategy of sorbitol feeding and low-temperature induction leads to high-level production of alkaline β-mannanase in Pichia pastoris.

A process for efficient production of an alkaline β-mannanases from Bacillus sp. N16-5 was established by heterologous expression using Pichia pastoris. A high producing strain was generated by removing the native β-mannanases signal peptide and increasing the copy number of the mature β-mannanases gene.

Understanding the effect of foreign gene dosage on the physiology of Pichia pastoris by transcriptional analysis of key genes.

Increased copy number of foreign gene can result in the alteration of normal metabolism in Pichia pastoris. To better understand the effect of foreign gene dosage on the cellular physiology of P. pastoris cells, comparative transcriptional analysis was performed among three P.

A systematical investigation on the genetic stability of multi-copy Pichia pastoris strains.

A battery of Pichia pastoris transformants, G1, G6, A2, A3, C3, carrying 1, 6, 12, 18 and 29 copies of porcine insulin precursor (PIP) gene, were employed to investigate the genetic stability of these multi-copy P. pastoris strains. Both G6 and C3 maintained their original copy numbers in serial culture without methanol induction for 35 generations.