Pyridoxal 5′-phosphate (PLP) functions as a coenzyme in many enzymatic processes, including decarboxylation, deamination, transamination, racemization, and others. Enzymes, requiring PLP, are commonly termed PLP-dependent enzymes, and they are widely involved in crucial cellular metabolic pathways in most of (if not all) living organisms. The chemical mechanisms for PLP-mediated reactions have been well elaborated and accepted with an emphasis on the pure chemical steps, but how the chemical steps are processed by enzymes, especially by functions of active site residues, are not fully elucidated. Furthermore, the specific mechanism of an enzyme in relation to the one for a similar class of enzymes seems scarcely described or discussed. This discussion aims to link the specific mechanism described for the individual enzyme to the same types of enzymes from different species with aminotransferases, decarboxylases, racemase, aldolase, cystathionine β-synthase, aromatic phenylacetaldehyde synthase, et al. as models. The structural factors that contribute to the reaction mechanisms, particularly active site residues critical for dictating the reaction specificity, are summarized in this review.

Bacteroidales are the most abundant order of bacteria in the healthy human gut and have the potential as a therapeutic agent. We constructed a pnCasBS-CBE system for base editing in the Bacteroides thetaiotaomicron to expand their genetic toolkit, which is able to efficiently convert a C:G to a T:A in the genome. As a functional proof-of-concept, we used the pnCasBS-CBE system to successfully introduce nonsynonymous mutation and stop codons to the genes involved in carbohydrate metabolism.The system also allowed for multiplexed gene editing with a single plasmid, enabling efficient editing of up to four genes in a single experiment. Furthermore, the pnCasBS-CBE editing system was validated and successfully applied in four other non-model gut Bacteroides species for genome editing. An unbiased genome-wide SNPs analysis indicated that the pnCasBS-CBE system showed high fidelity and applicability.Thus, this study provides a powerful clustered regularly interspaced short palindromic repeats (CRISPR)-mediated genome editing toolbox for functional genomic analysis in Bacteroidales.

Global aquaculture production has increased by 500% since the late 1980s to address the growing demand for aquatic proteins and decreasing fishery resources in natural aquatic ecosystems (FAO, 2016). More than 40% of global aquaculture production is carried out in earthen aquaculture ponds (Yuan et al., 2019). A total volume of 101 million tons of aquaculture was produced in 2014, and this volume is expected to continuously increase to 230 million tons in the next 20 years (FAO, 2014). Such huge increases in aquatic production depend heavily on