4'-Deoxypyridoxine disrupts vitamin B6 homeostasis in Escherichia coli K12 through combined inhibition of cumulative B6 uptake and PLP-dependent enzyme activity

Pyridoxal 5 '-phosphate (PLP) is the active form of vitamin B6 and a cofactor for many essential metabolic processes such as amino acid biosynthesis and one carbon metabolism. 4'-deoxypyridoxine (4dPN) is a long known B6 antimetabolite but its mechanism of action was not totally clear. By exploring different conditions in which PLP metabolism is affected in the model organism Escherichia coli K12, we showed that 4dPN cannot be used as a source of vitamin B6 as previously claimed and that it is toxic in several conditions where vitamin B6 homeostasis is affected, such as in a B6 auxotroph or in a mutant lacking the recently discovered PLP homeostasis gene, yggS. In addition, we found that 4dPN sensitivity is likely the result of multiple modes of toxicity, including inhibition of PLP-dependent enzyme activity by 4'-deoxypyridoxine phosphate (4dPNP) and inhibi-tion of cumulative pyridoxine (PN) uptake. These toxicities are largely dependent on the phosphorylation of 4dPN by pyridoxal kinase (PdxK)

Characterization of the Escherichia coli pyridoxal 5'-phosphate homeostasis protein (YggS): Role of lysine residues in PLP binding and protein stability

The pyridoxal 5'-phosphate (PLP) homeostasis protein (PLPHP) is a ubiquitous member of the COG0325 family with apparently no catalytic activity. Although the actual cellular role of this protein is unknown, it has been observed that mutations of the PLPHP encoding gene affect the activity of PLP-dependent enzymes, B6 vitamers and amino acid levels. Here we report a detailed characterization of the Escherichia coli ortholog of PLPHP (YggS) with respect to its PLP binding and transfer properties, stability, and structure. YggS binds PLP very tightly and is able to slowly transfer it to a model PLP-dependent enzyme, serine hydroxymethyltransferase. PLP binding to YggS elicits a conformational/flexibility change in the protein structure that is detectable in solution but not in crystals. We serendipitously discovered that the K36A variant of YggS, affecting the lysine residue that binds PLP at the active site, is able to bind PLP covalently. This observation led us to recognize that a number of lysine residues, located at the entrance of the active site, can replace Lys36 in its PLP binding role. These lysines form a cluster of charged residues that affect protein stability and conformation, playing an important role in PLP binding and possibly in YggS function

Screening and Brief Intervention for Underage Drinkers

In a 2007 report, the US Surgeon General called for health care professionals to renew efforts to reduce underage drinking. Focusing on the adolescent patient, this review provides health care professionals with recommendations for alcohol-related screening, brief intervention, and referral to treatment. MEDLINE and published reviews were used to identify relevant literature. Several brief screening methods have been shown to effectively identify underage drinkers likely to have alcohol use disorders. After diagnostic assessment when germane, the initial intervention typically focuses on education, motivation for change, and consideration of treatment options. Internet-accessible resources providing effective brief interventions are available, along with supplemental suggestions for parents. Recent changes in federal and commercial insurance reimbursement policies provide some fiscal support for these services, although rate increases and expanded applicability may be required to prompt the participation of many practitioners. Nevertheless, advances in clinical methods and progress on reimbursement policies have made screening and brief intervention for underage drinking more feasible in general health care practice

A roadmap for the functional annotation of protein families: a community perspective.

Funder: U.S. National Library of MedicineOver the last 25 years, biology has entered the genomic era and is becoming a science of 'big data'. Most interpretations of genomic analyses rely on accurate functional annotations of the proteins encoded by more than 500 000 genomes sequenced to date. By different estimates, only half the predicted sequenced proteins carry an accurate functional annotation, and this percentage varies drastically between different organismal lineages. Such a large gap in knowledge hampers all aspects of biological enterprise and, thereby, is standing in the way of genomic biology reaching its full potential. A brainstorming meeting to address this issue funded by the National Science Foundation was held during 3-4 February 2022. Bringing together data scientists, biocurators, computational biologists and experimentalists within the same venue allowed for a comprehensive assessment of the current state of functional annotations of protein families. Further, major issues that were obstructing the field were identified and discussed, which ultimately allowed for the proposal of solutions on how to move forward