Tag Archives: Enzymes

Scientists Found Superworms That Love Eating Styrofoam, And It Could Be a Good Thing

Packing material, disposable cutlery, CD cases: Polystyrene is among the most common forms of plastic, but recycling it isn’t easy and the vast majority ends up in landfills or finds its way to the oceans where it threatens marine life.

 

Scientists at Australia’s University of Queensland have now discovered that superworms – the larvae of Zophobas morio darkling beetles – are eager to dine on the substance, and their gut enzymes could hold the key to higher recycling rates.

Chris Rinke, who led a study that was published in the journal Microbial Genomics on Thursday, told AFP previous reports had shown that tiny waxworms and mealworms (which are also beetle larvae) had a good track record when it came to eating plastic, “so we hypothesized that the much larger superworms can eat even more.”

Superworms grow up to two inches (five centimeters) and are bred as a food source for reptiles and birds, or even for humans in countries such as Thailand and Mexico.

Rinke and his team fed superworms different diets over a three week period, with some given polystyrene foam, commonly known as styrofoam, some bran, and others not fed at all.

“We confirmed that superworms can survive on a sole polystyrene diet, and even gain a small amount of weight – compared to a starvation control group – which suggests that the worms can gain energy from eating polystyrene,” he said.

Polystyrene in the gut of a worm. (University of Queensland)

Although the polystyrene-reared superworms completed their life cycle, becoming pupae and then fully developed adult beetles, tests revealed a loss of microbial diversity in their guts and potential pathogens.

These findings suggested that while the bugs can survive on polystyrene, it is not a nutritious diet and impacts their health.

Next, the team used a technique called metagenomics to analyze the microbial gut community and find which gene-encoded enzymes were involved in degrading the plastic.

Bio-upcycling

One way to put the findings to use would be to provide superworms with food waste or agricultural bioproducts to consume alongside polystyrene.

“This could be a way to improve the health of the worms and to deal with the large amount of food waste in Western countries,” said Rinke.

 

But while breeding more worms for this purpose is possible, he envisages another route: creating recycling plants that mimic what the larvae do, which is to first shred the plastic in their mouths then digest it through bacterial enzymes.

“Ultimately, we want to take the superworms out of the equation,” he said, and he now plans more research aimed at finding the most efficient enzymes, then enhancing them further through enzyme engineering.

The breakdown products from that reaction could then be fed to other microbes to create high-value compounds, such as bioplastics, in what he hopes would become an economically viable “upcycling” approach.

© Agence France-Presse

 

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A COVID Diagnostic in Only 20 Minutes, Using Two CRISPR Enzymes

The reactions involved in the FIND-IT assay to detect infection with the SARS-CoV-2 virus. When the Cas13 enzyme (left) binds to its target RNA, it snips a molecule (orange and grey) to release an activator (orange) that supercharges the Csm6 nuclease (bottom center) to cleave and release fluorescent markers that light up (green) and signal the presence of viral RNA. Credit: Artwork courtesy of Margaret L. Liu, University of Chicago Pritzker School of Medicine

Frequent, rapid testing for

Tina Liu and Jennifer Doudna outside the IGI building on the day Doudna won the 2020 Nobel Prize in Chemistry. Credit: UC Berkeley photo by Brittany Hosea-Small

Several CRISPR-based assays have been authorized for emergency use by the Food and Drug Administration, but all require an initial step in which the viral RNA is amplified so that the detection signal — which involves release of a fluorescent molecule that glows under blue light — is bright enough to see. While this initial amplification increases the test’s sensitivity to a similar level as qRT-PCR, it also introduces steps that make the test more difficult to carry out outside of a laboratory.

The UC Berkeley-led team sought to reach a useful sensitivity and speed without sacrificing the simplicity of the assay.

“For point of care applications, you want to have a rapid response so that people can quickly know if they’re infected or not, before you get on a flight, for example, or go visit relatives,” said team leader Tina Liu, a research scientist in Doudna’s lab at the Innovative Genomics Institute (IGI), a CRISPR-focused center involving UC Berkeley and UC San Francisco scientists.

Aside from having an added step, another disadvantage of initial amplification is that, because it makes billions of copies of viral RNA, there is a greater chance of cross-contamination across patient samples. The new technique developed by the team flips this around and instead boosts the fluorescent signal, eliminating a major source of cross-contamination.

The amplification-free technique, which they term Fast Integrated Nuclease Detection In Tandem (FIND-IT), could enable quick and inexpensive diagnostic tests for many other infectious diseases.

“While we did start this project for the express purpose of impacting COVID-19, I think this particular technique could be applicable to more than just this pandemic because, ultimately, CRISPR is programable,” Liu said. “So, you could load the CRISPR enzyme with a sequence targeting flu virus or HIV virus or any type of RNA virus, and the system has the potential to work in the same way. This paper really establishes that this biochemistry is a simpler way to detect RNA and has the capability to detect that RNA in a sensitive and fast time frame that could be amenable for future applications in point of care diagnostics.”

The researchers are currently in the process of building such a diagnostic using FIND-IT, which would include steps to collect and process samples and to run the assay on a compact microfluidic device.

Employing tandem Cas proteins

To remove target amplification from the equation, the team employed a CRISPR enzyme — Cas13 — to first detect the viral RNA, and another type of Cas protein, called Csm6, to amplify the fluorescence signal.

Cas13 is a general purpose scissors for cutting RNA; once it binds to its target sequence, specified by a guide RNA, it is primed to cut a broad range of other RNA molecules. This target-triggered cutting activity can be harnessed to couple detection of a specific RNA sequence to release of a fluorescent reporter molecule. However, on its own, Cas13 can require hours to generate a detectable signal when very low amounts of target RNA are present.

Liu’s insight was to use Csm6 to amplify the effect of Cas13. Csm6 is a CRISPR enzyme that senses the presence of small rings of RNA and becomes activated to cut a broad range of RNA molecules in cells.

To boost Cas13 detection, she and her colleagues designed a specially engineered activator molecule that gets cut when Cas13 detects viral RNA. A fragment of this molecule can bind to and trigger Csm6 to cut and release a bright fluorescent molecule from a piece of RNA. Normally, the activator molecule is quickly broken down by Csm6, thus limiting the amount of fluorescent signal it can generate. Liu and her colleagues devised a way to chemically modify the activator so that it is protected from degradation and can supercharge Csm6 to repeatedly cut and release fluorescent molecules linked to RNA. This results in a sensitivity that is 100 times better than the original activator.

“When Cas13 gets activated, it cleaves this small activator, removing a segment that protects it,” Liu said. “Now that it’s liberated, it can activate lots of different molecules of that second enzyme, Csm6. And so, one target recognized by Cas13 doesn’t just lead to activation of its own RNA-cutting ability; it leads to the generation of many more active enzymes that can each then cleave even more fluorescent reporters.”

The team of researchers also incorporated an optimized combination of guide RNAs that enables more sensitive recognition of the viral RNA by Cas13. When this was combined with Csm6 and its activator, the team was able to detect down to 31 copies per microliter of (function(d, s, id){ var js, fjs = d.getElementsByTagName(s)[0]; if (d.getElementById(id)) return; js = d.createElement(s); js.id = id; js.src = "https://connect.facebook.net/en_US/sdk.js#xfbml=1&version=v2.6"; fjs.parentNode.insertBefore(js, fjs); }(document, 'script', 'facebook-jssdk'));

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