Search

Phenylalanine, as an essential aromatic amino acid, is not only needed for protein and vital molecules such as neurotransmitter and hormone synthesis but also a substrate for the biosynthesis of phenylpropanoids and various bioactive compounds. The metabolism of phenylalanine is dynamic and transitory, which would otherwise inhibit cell growth. Therefore, it is challenging and imperative to monitor intracellular phenylalanine bioavailability in real time, which has great significance for evaluating the effectiveness of introducing pathway-specific genetic modifications to enhance phenylalanine generation. In this study, we proposed a live-cell biomonitoring system to assess phenylalanine bioavailability in real time in cyanobacteria based on codon degeneracy and species-specific usage bias. The biomonitoring system was generated through genetic modification of phenylalanine codons in the chloramphenicol antibiotic resistance gene to wholly preferred and rare codons, in combination with an orthogonal constitutive promoter Trc to express these genes. Cyanobacterial cells equipped with a preferred codon-based gene showed a significant growth advantage over those with rare codons under antibiotic pressure, while the delayed growth caused by rare codon-based genes could be rescued by supplementing phenylalanine in the cultivation medium. Increasing intracellular phenylalanine bioavailability could promote rare codon-based gene containing cell growth to a similar level as wild-type strains harboring preferred codon-based gene, providing a live-cell visualized screening method to relatively define phenylalanine content from either random mutation libraries or pathway-specific engineering cyanobacterial chassis before conducting labor-intensive quantitative measurements., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Numerous plant secondary metabolites have remarkable impacts on both food supplements and pharmaceuticals for human health improvement. However, higher plants can only generate small amounts of these chemicals with specific temporal and spatial arrangements, which are unable to satisfy the expanding market demands. Cyanobacteria can directly utilize CO 2 , light energy, and inorganic nutrients to synthesize versatile plant-specific photosynthetic intermediates and organic compounds in large-scale photobioreactors with outstanding economic merit. Thus, they have been rapidly developed as a "green" chassis for the synthesis of bioproducts. Flavonoids, chemical compounds based on aromatic amino acids, are considered to be indispensable components in a variety of nutraceutical, pharmaceutical, and cosmetic applications. In contrast to heterotrophic metabolic engineering pioneers, such as yeast and Escherichia coli , information about the biosynthesis flavonoids and their derivatives is less comprehensive than that of their photosynthetic counterparts. Here, we review both benefits and challenges to promote cyanobacterial cell factories for flavonoid biosynthesis. With increasing concerns about global environmental issues and food security, we are confident that energy self-supporting cyanobacteria will attract increasing attention for the generation of different kinds of bioproducts. We hope that the work presented here will serve as an index and encourage more scientists to join in the relevant research area.

AbstractCyanobacteria, the only oxygenic photoautotrophs among prokaryotes, are developing as both carbon building blocks and energetic self-supported chassis for the generation of various bioproducts. However, one of the challenges to optimize it as a more sustainable platform is how to release intracellular bioproducts for an easier downstream biorefinery process. To date, the major method used for cyanobacterial cell lysis is based on mechanical force, which is energy-intensive and economically unsustainable. Phage-mediated bacterial cell lysis is species-specific and highly efficient and can be conducted under mild conditions; therefore, it has been intensively studied as a bacterial cell lysis weapon. In contrast to heterotrophic bacteria, biological cell lysis studies in cyanobacteria are lagging behind. In this study, we reviewed cyanobacterial cell envelope features that could affect cell strength and elicited a thorough presentation of the necessary phage lysin components for efficient cell lysis. We then summarized all bioengineering manipulated pipelines for lysin component optimization and further revealed the challenges for each intent-oriented application in cyanobacterial cell lysis. In addition to applied biotechnology usage, the significance of phage-mediated cyanobacterial cell lysis could also advance sophisticated biochemical studies and promote biocontrol of toxic cyanobacteria blooms. [ABSTRACT FROM AUTHOR]

Summary: Protein quality control pathways require AAA+ proteases, such as Clp and Lon. Lon protease maintains UmuD, an important component of the error‐prone DNA repair polymerase (Pol V), at very low levels in E. coli. Most members of the phylum Cyanobacteria lack Lon (including the model cyanobacterium, Synechocystis sp. PCC6803), so maintenance of UmuD at low levels must employ different proteases. We demonstrate that the first 19 residues from the N‐terminus of UmuD (Sug1‐19) fused to a reporter protein are adequate to trigger complete proteolysis and that mutation of a single leucine residue (L6) to aspartic acid inhibits proteolysis. This process appears to follow the N‐end rule and is mediated by ClpA/P protease and the ClpS adaptor. Additionally, mutations of arginine residues in the Sug1‐19 tag suggest that the ClpX/P pathway also plays a role in proteolysis. We propose that there is a dual degron at the N‐terminus of the UmuD protein in Synechocystis sp. PCC6803, which is distinct from the degron required for degradation of UmuD in E. coli. The use of two proteolysis pathways to tune levels of UmuD might reflect how a photosynthetic organism responds to multiple environmental stressors. [ABSTRACT FROM AUTHOR]