Nucleic-acid detection via isothermal amplification and collateral cleavage of reporter molecules by CRISPR-associated enzymes is a promising alternative to quantitative polymerase chain reaction (qPCR). Here, we report the clinical validation of the SHERLOCK (specific high-sensitivity enzymatic reporter unlocking) assay using the enzyme Cas13a from Leptotrichia wadei for the detection of SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) -the virus that causes COVID-19 (coronavirus disease 2019) -in 154 nasopharyngeal and throat swab samples collected at Siriraj Hospital, Thailand. Within a detection limit of 42 RNA copies per

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Phenolic acids are abundant biomass feedstock that can be derived from the processing of lignin or other byproducts from agro-industrial waste. Although phenolic acids such as p-hydroxybenzoic acid, p-coumaric acid, caffeic acid, vanillic acid, cinnamic acid, gallic acid, syringic acid, and ferulic acid can be used directly in various applications, their value can be significantly increased when they are further modified to high value-added compounds. This review summarizes and discusses the new advances in cell-free and whole-cell biocatalysis technologies for reactions important for conversion of phenolic acids including esterification, decarboxylation, amination, halogenation, hydroxylation, and ring-breakage reactions. The products of these reactions are useful for the pharmaceutical, cosmetic, food, fragrance, and polymer industries. Production of phenolic acids is sustainable, and these processes for their biotransformation are clean technologies that do not produce toxic waste and use less energy than conventional physical and chemical methods. Thus, biotransformation of phenolic acids provides an economically viable and sustainable means for producing useful materials for society.

The putative gene of Plasmodium vivax serine hydroxymethyltransferase (PvSHMT; EC 2.1.2.1) was cloned and expressed in Escherichia coli. The purified enzyme was shown to be a dimeric protein with a monomeric molecular mass of 49 kDa. PvSHMT has a maximum absorption peak at 422 nm with a molar absorption coefficient of 6370 m−1·cm−1. The Kd for binding of the enzyme and pyridoxal‐5‐phosphate was 0.14 ± 0.01 μm. An alternative assay for measuring the tetrahydrofolate‐dependent SHMT activity based on the coupled reaction with 5,10‐methylenetetrahydrofolate reductase (EC 1.5.1.20) from E. coli was developed. PvSHMT uses a ternary‐complex mechanism with a kcat value of 0.98 ± 0.06 s−1 and Km values of 0.18 ± 0.03 and 0.14 ± 0.02  mm for l‐serine and tetrahydrofolate, respectively. The optimum pH of the SHMT reaction was 8.0 and an Arrhenius’s plot showed a transition temperature of 19 °C. Besides l‐serine, PvSHMT forms an external aldimine complex with d‐serine, l‐alanine, l‐threonine and glycine. PvSHMT also catalyzes the tetrahydrofolate‐independent retro‐aldol cleavage of 3‐hydroxy amino acids. Although l‐serine is a physiological substrate for SHMT in the tetrahydrofolate‐dependent reaction, PvSHMT can also use d‐serine. In the absence of tetrahydrofolate at high pH, PvSHMT forms an enzyme–quinonoid complex with d‐serine, but not with l‐serine, whereas SHMT from rabbit liver was reported to form an enzyme–quinonoid complex with l‐serine. The substrate specificity difference between PvSHMT and the mammalian enzyme indicates the dissimilarity between their active sites, which could be exploited for the development of specific inhibitors against PvSHMT.

Plasmodiumparasites, the causative agent of malaria, rely heavily onde novofolate biosynthesis, and the enzymes in this pathway have therefore been explored extensively for antimalarial development. Serine hydroxymethyltransferase (SHMT) fromPlasmodiumspp., an enzyme involved in folate recycling and dTMP synthesis, has been shown to catalyze the conversion of L- and D-serine to glycine (Gly) in a THF-dependent reaction, the mechanism of which is not yet fully understood. Here, the crystal structures ofP. vivaxSHMT (PvSHMT) in a binary complex with L-serine and in a ternary complex with D-serine (D-Ser) and (6R)-5-formyltetrahydrofolate (5FTHF) provide clues to the mechanism underlying the control of enzyme activity. 5FTHF in the ternary-complex structure was found in the 6Rform, thus differing from the previously reported structures of SHMT–Gly–(6S)-5FTHF from other organisms. This suggested that the presence of D-Ser in the active site can alter the folate-binding specificity. Investigation of binding in the presence of D-Ser and the (6R)- or (6S)-5FTHF enantiomers indicated that both forms of 5FTHF can bind to the enzyme but that only (6S)-5FTHF gives rise to a quinonoid intermediate. Likewise, a large surface area with a highly positively charged electrostatic potential surrounding thePvSHMT folate pocket suggested a preference for a polyglutamated folate substrate similar to the mammalian SHMTs. Furthermore, as inP. falciparumSHMT, a redox switch created from a cysteine pair (Cys125–Cys364) was observed. Overall, these results assert the importance of features such as stereoselectivity and redox status for control of the activity and specificity ofPvSHMT.