"Aquaplastic" Made From Bacteria - 2021.03.22

Plus: All the other synthetic biology research this week.

☀️ Good morning.

Some paintings become famous because, being durable, they are viewed by successive generations, in each of which are likely to be found a few appreciative eyes. I know a painting so evanescent that it is seldom viewed at all, except by some wandering deer. It is a river who wields the brush, and it is the same river who, before I can bring my friends to view his work, erases it forever.

—Aldo Leopold, in “A Sand County Almanac.”

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Water Dissolvable Plastics from Engineered Biofilms: Biomaterials are already amazing, but synthetic biology has pushed their properties to new heights: cellulose pellicles with embedded yeast, mushroom leather, synthetic spider silk. And now … aquaplastic.

For a study in Nature Chemical Biology, titled “Water-processable, biodegradable and coatable aquaplastic from engineered biofilms,” researchers from the Joshi lab at Harvard University created a material, produced from living cells, that can be healed or welded together using water.

Plastics take a long time to degrade, and so bio-based alternatives are desired. The researchers first engineered E. coli bacteria to produce curli fibers—long protein chains—and excrete them into their environment. The researchers then filtered the curli proteins from the cells, producing a gel that “can be cast onto surfaces or molds to create plastic films.”

Unlike traditional plastics, these “aquaplastics” don’t need heat to set; they dry and harden at room temperature. They are also flexible, and can be twisted and bent into different positions without tearing or breaking. The researchers put the aquaplastic through a wide range of tests, measuring their hardness, yield strength, and other material properties. They were similar to conventional plastics.

The aquaplastics are also biodegradable. The curli-based plastic lost 89 percent of its mass over 45 days when exposed to “a mixed culture of microorganisms obtained from the primary effluent of a wastewater treatment plant.” That is an improvement over nanocellulose—a highly biodegradable material—which lost 85 percent of its mass in that same time span.

In a final batch of experiments, the researchers used water to heal scratches and tears in the aquaplastic. Scratches “could be removed completely by spraying a few microliters of water onto the scratch and allowing it to air-dry for 2min.” A similar feat was achieved for fully cut material.

Why It Matters: Plastic pollution is an urgent, global problem. Cheap, biodegradable alternatives that are easy to produce are needed. This study shows that a templatable, coatable “plastic” can be made from living, engineered cells. This material could be used for packaging in the future; the authors are already working to increase the yield of aquaplastic production from living cells, which would decrease costs and make this a more viable plastic alternative.


Competition in CRISPR Circuits: The “deactivated” form of the gene-editing protein Cas9—called dCas9—can bind to target DNA, but cannot make cuts. dCas9 can repress any target gene in a cell just by changing the sequence of its guide RNA. This makes it useful for building genetic circuits.

Genetic circuits built with CRISPR, though, have a major drawback: as more bells and whistles are added to the circuit, so too must the number of guide RNAs increase. With more guide RNAs floating around the cell, they begin to jockey for dCas9’s attention. This competition makes larger CRISPR-based genetic circuits, with more components, unreliable.

For a new study, in Nature Communications, a team led by Domitilla Del Vecchio at MIT created a “dCas9 concentration regulator.” It is a genetic control system that can tune the amount of dCas9 protein in a cell based on how many different guide RNAs are present.

The researchers created a “feedback regulator” that increases the level of dCas9 when there are more guide RNAs in the cell, and decreases dCas9’s level when there are fewer. To do that, they continuously express a guide RNA (let’s call it guide-0) that binds to dCas9 and orders the protein to repress its own promoter.

Since guide-0 is produced at a steady level, dCas9 represses itself at a fixed level. But as more guide RNAs of different sequences are added to the cell, they compete for dCas9’s attention, and guide-0 is less able to bind dCas9; the “repression” of dCas9’s promoter is then reduced and the amount of dCas9 increases. It’s a simple feedback system that works surprisingly well.

The authors use this approach to build a variety of genetic circuits with up to four total guide RNAs.

Del Vecchio wrote a blog post about the paper, and Pablo Cárdenas, a co-author, tweeted a great thread about it as well.

Why It Matters: CRISPR-based genetic circuits are easy to program and build. dCas9 can repress or activate any target gene in a cell. But it has been challenging to build complicated circuits using this approach because of the competition between guide RNAs. This study is a clever, useful approach for regulating dCas9 levels in response to additional guide RNAs. It could help bioengineers build larger, more complicated CRISPR-based genetic circuits.


30-Minute Coronavirus Test: Many research groups have developed CRISPR-based diagnostic tests for SARS-CoV-2 in the last year. Zhang, Abudayyeh and Gootenberg released a protocol and published a paper in the New England Journal of Medicine, last September, showing how a Cas12a protein, from Alicyclobacillus acidiphilus, could be used to detect SARS-CoV-2 RNA directly from nasal swabs. Their diagnostic test could detect just 100 copies of viral genome in a sample, in a process that took just 45 minutes. Their test had a sensitivity of 93.1 percent and a specificity of 98.5 percent. Other groups published similar studies.

Those early CRISPR diagnostic papers were not necessarily useful for detecting variants of the novel coronavirus, however.

For a new study in Nature Communications, titled “An engineered CRISPR-Cas12a variant and DNA-RNA hybrid guides enable robust and rapid COVID-19 testing,” a team led by Meng How Tan at the Nanyang Technological University, in Singapore, engineered a Cas12a protein from Acidaminococcus that can “tolerate single nucleotide variations in the target sites” of SARS-CoV-2 variants. The engineered Cas12a protein was made by changing just three amino acids—E174R/S542R/K548R. Despite its “promiscuity,” it maintained a high specificity for SARS-CoV-2 genomes.

By further modifying the guide RNAs, the authors also reduced off-target recognition of the engineered Cas12a protein. Their test could be completed in 30 minutes, and they were able to detect just 40 copies of a SARS-CoV-2 genome in one milliliter of a sample, straight from a nasal swab. Their test could detect a wide array of SARS-CoV-2 variants, too.

Why It Matters: While there are a lot of CRISPR-based tests that have been developed for COVID-19, it is unknown whether they can be used to detect a wide range of viral variants. The main advancement of this new study is the engineering of a Cas12a variant that can tolerate “single nucleotide variations” between clinical samples. This new enzyme could possibly be used to detect SARS-CoV-2 variants that have not emerged yet.


For a new paper in eLife, researchers found that traffic jams within dense, microbial spaces can lead to antibiotic-resistant biofilms. Check out the great video in this tweet from the lead author. 👇


🧫 Other Studies Published This Week

Am I missing coverage on a certain topic? Please leave a comment on this post.

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Artificial Life

  • Engineering motile aqueous phase-separated droplets via liposome stabilisation. Nature Communications. Open Access. Link

Biomaterials

  • A mussel-inspired film for adhesion to wet buccal tissue and efficient buccal drug delivery. Nature Communications. Open Access. Link

Biosensors

  • Programming cell-free biosensors with DNA strand displacement circuits. bioRxiv (preprint). Link

  • AL-PHA beads: Bioplastic-based protease biosensors for global health applications. Materials Today. Open Access. Link

Circuits

  • Harnessing the central dogma for stringent multi-level control of gene expression. Nature Communications. Open Access. Link

  • 2D printed multicellular devices performing digital and analogue computation. Nature Communications. Open Access. Link

Genetic Engineering & Control

  • Discovery and engineering of small SlugCas9 with broad targeting range and high specificity and activity. Nucleic Acids Research. Open Access. Link

  • Exploiting heterologous and endogenous CRISPR‐Cas systems for genome editing in the probiotic Clostridium butyricum. Biotechnology and Bioengineering. Link

  • Antisense RNA interference-enhanced CRISPR/Cas9 base editing method for improving base editing efficiency in Streptomyces lividans 66. ACS Synthetic Biology. Link

  • Harnessing the power of directed evolution to improve genome editing systems (Review). Current Opinion in Chemical Biology. Link

Medicine and Diagnostics

  • Ex utero mouse embryogenesis from pre-gastrulation to late organogenesis. Nature. Link

  • A pocket-sized device automates multiplexed point-of-care RNA testing for rapid screening of infectious pathogens. Biosensors and Bioelectronics. Link

  • Targeting loss of heterozygosity for cancer-specific immunotherapy. PNAS. Open Access. Link

  • Amelioration of hemophilia B through CRISPR/Cas9 induced homology-independent targeted integration. bioRxiv (preprint). Link

  • Site-specific antigen-adjuvant conjugation using cell-free protein synthesis enhances antigen presentation and CD8+ T-cell response. Scientific Reports. Open Access. Link

Metabolic Engineering

  • Control and regulation of acetate overflow in Escherichia coli. eLife. Open Access. Link

  • Auxiliary module promotes the synthesis of carboxysomes in E. coli to achieve high-efficiency CO2 assimilation. ACS Synthetic Biology. Link

  • Computational design and analysis of modular cells for large libraries of exchangeable product synthesis modules. bioRxiv (preprint). Link

  • In vivo and in vitro reconstitution of unique key steps in cystobactamid antibiotic biosynthesis. Nature Communications. Open Access. Link

  • An aldolase-based new pathway for bioconversion of formaldehyde and ethanol into 1,3-propanediol in Escherichia coli. ACS Synthetic Biology. Link

  • A cell factory of a fungicolous fungus Calcarisporiumarbuscula for efficient production of natural products. ACS Synthetic Biology. Link

  • Multiproduct microalgae biorefineries mediated by ionic liquids. Trends in Biotechnology. Open Access. Link

  • Simultaneous improvement of limonene production and tolerance in Yarrowia lipolytica through tolerance engineering and evolutionary engineering. ACS Synthetic Biology. Link

  • Identification and characterization of the mitochondrial replication origin for stable and episomal expression in Yarrowia lipolytica. ACS Synthetic Biology. Link

New Technology

  • A chemogenetic platform for controlling plasma membrane signaling and synthetic signal oscillation. bioRxiv (preprint). Link

Plants

  • Plant synthetic biology for producing potent phyto-antimicrobials to combat antimicrobial resistance. Biotechnology Advances. Link

Protein Engineering

  • Single-component, self-assembling, protein nanoparticles presenting the receptor binding domain and stabilized spike as SARS-CoV-2 vaccine candidates. Science Advances. Open Access. Link

Systems Biology & Modelling

  • Estimating maximal microbial growth rates from cultures, metagenomes, and single cells via codon usage patterns. PNAS. Open Access. Link

  • Programmable pattern formation in cellular systems with local signaling. bioRxiv (preprint). Link

  • The circadian oscillator analysed at the single‐transcript level. Molecular Systems Biology. Open Access. Link

  • Local genetic context shapes the function of a gene regulatory network. eLife. Open Access. Link

  • Quantifying absolute gene expression profiles reveals distinct regulation of central carbon metabolism genes in yeast. eLife. Open Access. Link

  • Scaling down the microbial loop: data-driven modelling of growth interactions in a diatom-bacterium co-culture. bioRxiv (preprint). Link

  • Growth-rate dependent and nutrient-specific gene expression resource allocation in fission yeast. bioRxiv (preprint). Link

  • Accurate reconstruction of dynamic gene expression and growth rate profiles from noisy measurements. bioRxiv (preprint). Link

Miscellaneous Topics

  • Processive RNA polymerization and promoter recognition in an RNA World. Science. Link

  • Biotechnology and biosafety policy at OECD: Future trends. Trends in Biotechnology. Open Access. Link

Until next time,

— Niko

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