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Surface-enhanced Raman spectroscopy (SERS) is a powerful optical sensing technique used in various applications, including medicine, microbiology, and environmental analysis. Planar SERS substrates are of particular interest due to their ease of integration in lab-on-chips and better reproducibility compared to colloidal SERS. The performance of these SERS substrates is quantified using metrics such as enhancement factor, sensitivity, and reproducibility. However, there is yet to be a consensus on how to practically compare and interpret such metrics in publications and experiments. These performance metrics are strongly influenced by the nanostructures’ material, architecture, element sizes, as well as the circumstances surrounding the experiments. Understanding the effect of these characteristics on the SERS substrates’ performance could not only enable a better performance but also direct their development for different applications. Thus, we prepared a planar SERS-substrate characterization methodology to explore the correlation between the nanostructures’ physical characteristics and the performance metrics through coordinate-transformed spectroscopic measurements over structure-characterized areas. Seven commercial SERS substrates, with various surface architectures fabricated using different fabrication technologies, were studied using this benchmarking methodology. The results demonstrated how this methodology can indicate a SERS substrate’s suitability for a specific application, thus, guiding the substrate’s further adaptations or development.

Frameshifts can be caused by specific combinations of tRNA and mRNA. The wildtype AGC-decoding E. coli tRNA has been shown to induce -1 ribosomal frameshifting on GCA alanine codons, and proposed to read a two-base codon instead of a canonical triplet. However, it has remained unclear whether this type of non-cognate decoding can be accommodated by the ribosome. Here, we perform single-particle cryo-EM reconstructions on E. coli 70S ribosomes with the frameshift-inducing tRNA bound to the non-cognate GCA codon or the cognate AGC codon in the ribosomal A site. The structures demonstrate that doublet decoding is made possible when A1493, the conserved monitoring base in 16S rRNA, mimics a first codon base, forming a Hoogsteen base pair with U36 from the anticodon and stacking with the mRNA. This interaction pushes the first two bases of the A-site codon in position for base pairing with C35 and G34 of the anticodon.

Here, the structure of the 30S ribosomal subunit and the 70S ribosome in complex with a messenger RNA with pseudouridine in the place of uridine reveals unexpected base pairing. Surprising base pairs fool ribosome When messenger RNA is translated into protein, the end of the protein-coding sequence is indicated by a three-base stop codon. Stop codons do not code for an amino acid, but recently it was shown that changing the first base to a pseudouridine (Ψ, the C-glycoside isomer of the nucleoside uridine) allows the incorporation of an amino acid, such that translation could continue past the stop codon. Venki Ramakrishnan and colleagues have determined the structure of the 30S ribosomal subunit in complex with an mRNA with ΨAG at the A site and a portion of serine transfer RNA. The structure reveals unexpected purine–purine base pairing at the first position of the codon and unusual pairing at the second and third positions. This study provides additional evidence for plasticity in the decoding centre of the ribosome. During normal translation, the binding of a release factor to one of the three stop codons (UGA, UAA or UAG) results in the termination of protein synthesis. However, modification of the initial uridine to a pseudouridine (Ψ) allows efficient recognition and read-through of these stop codons by a transfer RNA (tRNA), although it requires the formation of two normally forbidden purine–purine base pairs 1 . Here we determined the crystal structure at 3.1 Å resolution of the 30S ribosomal subunit in complex with the anticodon stem loop of tRNA Ser bound to the ΨAG stop codon in the A site. The ΨA base pair at the first position is accompanied by the formation of purine–purine base pairs at the second and third positions of the codon, which show an unusual Watson–Crick/Hoogsteen geometry. The structure shows a previously unsuspected ability of the ribosomal decoding centre to accommodate non-canonical base pairs.

The Ser/Leu-swapped genetic code can act as a genetic firewall, mitigating biohazard risks arising from horizontal gene transfer in genetically modified organisms. Our prior work demonstrated the orthogonality of this swapped code to the standard genetic code using a cell-free translation system comprised of 21 in vitro transcribed tRNAs. In this study, to advance this system for protein engineering, we introduce a natural / in vitro transcribed-hybrid tRNA set. This set combines natural tRNAs from Escherichia coli (excluding Ser, Leu, and Tyr) and in vitro transcribed tRNAs, encompassing anticodon-swapped tRNA Ser GAG and tRNA Leu GGA . This approach reduces the number of in vitro transcribed tRNAs required from 21 to only 4. In this optimized system, the production of a model protein, superfolder green fluorescent protein, increases to 3.5-fold. With this hybrid tRNA set, the Ser/Leu-swapped cell-free translation system will stand as a potent tool for protein production with reduced biohazard concerns in future biological endeavors. The use of orthogonal genetic code can help to prevent the escape of hazardous genes through horizontal gene transfer. Here, the authors develop a cell-free translation system with the Ser/Leu-swapped genetic code using a hybrid tRNA set and show its application in enhancing the production of superfolder GFP.

Coleoptera is the most diverse group of insects with over 360,000 described species divided into four suborders: Adephaga, Archostemata, Myxophaga, and Polyphaga. In this study, we present six new complete mitochondrial genome (mtgenome) descriptions, including a representative of each suborder, and analyze the evolution of mtgenomes from a comparative framework using all available coleopteran mtgenomes. We propose a modification of atypical cox1 start codons based on sequence alignment to better reflect the conservation observed across species as well as findings of TTG start codons in other genes. We also analyze tRNA-Ser(AGN) anticodons, usually GCU in arthropods, and report a conserved UCU anticodon as a possible synapomorphy across Polyphaga. We further analyze the secondary structure of tRNA-Ser(AGN) and present a consensus structure and an updated covariance model that allows tRNAscan-SE (via the COVE software package) to locate and fold these atypical tRNAs with much greater consistency. We also report secondary structure predictions for both rRNA genes based on conserved stems. All six species of beetle have the same gene order as the ancestral insect. We report noncoding DNA regions, including a small gap region of about 20 bp between tRNA-Ser(UCN) and nad1 that is present in all six genomes, and present results of a base composition analysis. [PUBLICATION ABSTRACT]

Exosomes, carrying distinctive biomolecules reflective of their parent cell's status and origin, show promise as liquid biopsy biomarkers for cancer diagnosis. However, their clinical translation remains challenging due to their relatively low concentration in body fluids. Surface-Enhanced Raman spectroscopy (SERS) has recently gained significant attention as a label-free and sensitive technique for exosome analysis. This review explores label-free SERS for exosome detection, covering exosome isolation and characterization methods, advancements in SERS substrates, and fingerprint analysis techniques using machine learning. Furthermore, we emphasize the challenges and offer insights into the future prospects of SERS-based exosome analysis to enhance cancer diagnosis.

Selenium is a fascinating element that has a long history, most of which documents it as a deleterious element to health. In more recent years, selenium has been found to be an essential element in the diet of humans, all other mammals, and many other life forms. It has many health benefits that include, for example, roles in preventing heart disease and certain forms of cancer, slowing AIDS progression in HIV patients, supporting male reproduction, inhibiting viral expression, and boosting the immune system, and it also plays essential roles in mammalian development. Elucidating the molecular biology of selenium over the past 40 years generated an entirely new field of science which encompassed the many novel features of selenium. These features were (1) how this element makes its way into protein as the 21st amino acid in the genetic code, selenocysteine (Sec); (2) the vast amount of machinery dedicated to synthesizing Sec uniquely on its tRNA; (3) the incorporation of Sec into protein; and (4) the roles of the resulting Sec-containing proteins (selenoproteins) in health and development. One of the research areas receiving the most attention regarding selenium in health has been its role in cancer prevention, but further research has also exposed the role of this element as a facilitator of various maladies, including cancer.

Clusterin plays an important role in carcinogenesis and serves as an important diagnostic biomarker of various clinical conditions. This work describes an application of a surface-enhanced Raman scattering (SERS)-based immunoassay using Al foil substrate that has the potential for the detection of clusterin. We first optimized the parameters of the assay using anti-human IgG/human IgG (hIgG) as a model antibody/antigen system using various substrates based on Au film, Si, Al tape and Al foil. Among the tested substrates, Al foil exhibited better performance, when assay of human IgG on Al foil demonstrated a detection limit of 2 pM and a semi logarithmic trend range from 10 pM to 1000 pM. Afterwards, the same SERS immunoassay method was implemented for detection of clusterin and resulted in a good semi-logarithmic calibration line with a high R2 value of 0.99, which was obtained in the range from 1 ng/mL to 1000 ng/mL. The low detection limit for clusterin antigen was found to be 3 ng/mL, which is better than most LODs for clusterin reported in the literature and also nearly 4 orders of magnitude lower than possible concentrations of clusterin in human blood. Moreover, the assay requires a relatively low volume of sample (10 μL). Overall, the assay performance demonstrates the significant potential of SERS on Al foil as a low-cost/high-availability substrate for sensing and biosensing, including detection of cancer biomarkers.

Surface‐enhanced Raman scattering (SERS) is a powerful sensing technique. However, the employment of SERS sensors in practical applications is hindered by high fabrication costs from processes with limited scalability, poor batch‐to‐batch reproducibility, substrate stability, and uniformity. Here, highly scalable and reproducible flame aerosol technology is employed to rapidly self‐assemble uniform SERS sensing films. Plasmonic Ag nanoparticles are deposited on substrates as nanoaggregates with fine control of their interparticle distance. The interparticle distance is tuned by adding a dielectric spacer during nanoparticle synthesis that separates the individual Ag nanoparticles within each nanoaggregate. The dielectric spacer thickness dictates the plasmonic coupling extinction of the deposited nanoaggregates and finely tunes the Raman hotspots. By systematically studying the optical and morphological properties of the developed SERS surfaces, structure–performance relationships are established and the optimal hot‐spots occur for interparticle distance of 1 to 1.5 nm among the individual Ag nanoparticles, as also validated by computational modeling, are identified for the highest signal enhancement of a molecular Raman reporter. Finally, the superior stability and batch‐to‐batch reproducibility of the developed SERS sensors are demonstrated and their potential with a proof‐of‐concept practical application in food‐safety diagnostics for pesticide detection on fruit surfaces is explored. Robust surface‐enhanced Raman scattering (SERS) sensing surfaces are fabricated using one‐step flame nanoparticle deposition. The sensing surfaces exhibit superior stability and high batch‐to‐batch reproducibility, highlighting their potential in practical (bio)chemical sensing. The detection of pesticides on fruit surfaces demonstrates a proof‐of‐concept practical application in food safety diagnostics at the point of consumption.

The red swamp crayfish (Procambarus clarkii) is a widely distributed invasive species that is listed in the Delivering Alien Invasive Species Inventory for Europe. Native to North America, it has been introduced to numerous regions, such as the Canary Islands, Spain. Previous studies have confirmed the role of this crayfish in the maintenance of several foodborne pathogenic bacteria. Therefore, the aim of this study was to analyze the main zoonotic bacterial and parasitic pathogens present in a P. clarkii population introduced to the island of Tenerife, Canary Islands, and to assess the potential risk to public health and native fauna. A total of 22 crayfish from Tenerife were analyzed using Biofire FilmArray Gastrointestinal Panels and culture–PCR methods. The results show the presence of Plesiomonas shigelloides, Shigella/enteroinvasive Escherichia coli, enteropathogenic Escherichia coli, Salmonella ser. Enteritidis, Salmonella ser. Typhimurium, and Salmonella ser. Typhi. These results demonstrate the presence of a variety of pathogenic bacteria in the red swamp crayfish in Tenerife that represent a significant concern in terms of public health and conservation. Implementing educational campaigns to inform the community about the risks associated with handling and consuming contaminated crayfish, as well as initiatives for the restoration of the contaminated ecosystem, are necessary to prevent the transmission of the foodborne pathogens.