Regioselective preparation of 5-hydroxypropranolol and 4′-hydroxydiclofenac with a fungal peroxygenase

An extracellular peroxygenase of Agrocybe aegerita catalyzed the H₂O₂-dependent hydroxylation of the multi-function beta-adrenergic blocker propranolol (1-naphthalen-1-yloxy-3-(propan-2-ylamino)propan-2-ol) and the non-steroidal anti-inflammatory drug diclofenac (2-[2-[(2,6-dichlorophenyl)amino]phenyl]acetic acid) to give the human drug metabolites 5-hydroxypropranolol (5-OHP) and 4′-hydroxydiclofenac (4′-OHD). The reactions proceeded regioselectively with high isomeric purity and gave the desired 5-OHP and 4′-OHD in yields up to 20% and 65%, respectively. ¹⁸O-labeling experiments showed that the phenolic hydroxyl groups in 5-OHP and 4′-OHD originated from H₂O₂, which establishes that the reaction is mechanistically a peroxygenation. Our results raise the possibility that fungal peroxygenases may be useful for versatile, cost-effective, and scalable syntheses of drug metabolites.     

Crosstalk between environmental light and internal time in humans

Daily exposure to environmental light is the most important zeitgeber in humans, and all studied characteristics of light pattern (timing, intensity, rate of change, duration, and spectrum) influence the circadian system. However, and due to lack of current studies on environmental light exposure and its influence on the circadian system, the aim of this work is to determine the characteristics of a naturalistic regimen of light exposure and its relationship with the functioning of the human circadian system. Eighty-eight undergraduate students (18-23 yrs) were recruited in Murcia, Spain (latitude 38°01'N) to record wrist temperature (WT), light exposure, and sleep for 1 wk under free-living conditions. Light-exposure timing, rate of change, regularity, intensity, and contrast were calculated, and their effects on the sleep pattern and WT rhythm were then analyzed. In general, higher values for interdaily stability, relative amplitude, mean morning light, and light quality index (LQI) correlated with higher interdaily stability and relative amplitude, and phase advance in sleep plus greater stability in WT and phase advance of the WT circadian rhythm. On the other hand, a higher fragmentation of the light-exposure rhythm was associated with more fragmented sleep. Naturalistic studies using 24-h ambulatory light monitoring provide essential information about the main circadian system input, necessary for maintaining healthy circadian tuning. Correcting light-exposure patterns accordingly may help prevent or even reverse health problems associated with circadian disruption.     

Oligosaccharyltransferase isoforms that contain different catalytic STT3 subunits have distinct enzymatic properties

Oligosaccharyltransferase (OST) is an integral membrane protein that catalyzes N-linked glycosylation of nascent proteins in the lumen of the endoplasmic reticulum. Although the yeast OST is an octamer assembled from nonhomologous subunits (Ost1p, Ost2p, Ost3p/Ost6p, Ost4p, Ost5p, Wbp1p, Swp1p, and Stt3p), the composition of the vertebrate OST was less well defined. The roles of specific OST subunits remained enigmatic. Here we show that genomes of most multicellular eukaryotes encode two homologs of Stt3p and mammals express two homologs of Ost3p. The Stt3p and Ost3p homologs are assembled together with the previously described mammalian OST subunits (ribophorins I and II, OST48, and DAD1) into complexes that differ significantly in enzymatic activity. Tissue and cell type-specific differences in expression of the Stt3p homologs suggest that the enzymatic properties of oligosaccharyltransferase are regulated in eukaryotes to respond to alterations in glycoprotein flux through the secretory pathway and may contribute to tissue-specific glycan heterogeneity.     

Identification of a novel protein promoting the colonization and survival of Finegoldia magna, a bacterial commensal and opportunistic pathogen

Anaerobic bacteria dominate the human normal microbiota, but strikingly little is known about these commensals. Finegoldia magna is a Gram-positive anaerobe found in the skin and at other non-sterile body surfaces, but it is also an opportunistic pathogen. This study describes a novel protein designated FAF (F. magna adhesion factor) and expressed by more than 90% of F. magna isolates. The protein is present in substantial quantities at the F. magna surface but is also released from the surface. FAF forms large protein aggregates in solution and surface-associated FAF causes bacterial clumping. In skin F. magna bacteria were localized to the epidermis, where they adhere to basement membranes. FAF was found to mediate this adhesion via interactions with BM-40, a basement membrane protein. The biological significance of FAF is further underlined by the observation that it blocks the activity of LL-37, a major human antibacterial peptide. Altogether, the data demonstrate that FAF plays an important role in colonization and survival of F. magna in the human host.     

Sex roles and mutual mate choice matter during mate sampling

The roles of females and males in mating competition and mate choice have lately proven more variable, between and within species, than previously thought. In nature, mating competition occurs during mate search and is expected to be regulated by the numbers of potential mates and same-sex competitors. Here, we present the first study to test how a temporal change in sex roles affects mating competition and mate choice during mate sampling. Our model system (the marine fish Gobiusculus flavescens) is uniquely suitable because of its change in sex roles, from conventional to reversed, over the breeding season. As predicted from sex role theory, courtship was typically initiated by males and terminated by females early in the breeding season. The opposite pattern was observed late in the season, at which time several females often simultaneously courted the same male. Mate-searching females visited more males early than late in the breeding season. Our study shows that mutual mate choice and mating competition can have profound effects on female and male behavior. Future work needs to consider the dynamic nature of mating competition and mate choice if we aim to fully understand sexual selection in the wild.     

Perforin Promotes Amyloid Beta Internalisation in Neurons

Studies on the mechanisms of neuronal amyloid-β (Aβ) internalisation are crucial for understanding the neuropathological progression of Alzheimer’s disease (AD). We here investigated how extracellular Aβ peptides are internalised and focused on three different pathways: (i) via endocytic mechanisms, (ii) via the receptor for advanced glycation end products (RAGE) and (iii) via the pore-forming protein perforin. Both Aβ₄₀ and Aβ₄₂ were internalised in retinoic acid differentiated neuroblastoma (RA-SH-SY5Y) cells. A higher concentration was required for Aβ₄₀ (250 nM) compared with Aβ₄₂ (100 nM). The internalised Aβ₄₀ showed a dot-like pattern of distribution whereas Aβ₄₂ accumulated in larger and distinct formations. By confocal microscopy, we showed that Aβ₄₀ and Aβ₄₂ co-localised with mitochondria, endoplasmic reticulum (ER) and lysosomes. Aβ treatment of human primary cortical neurons (hPCN) confirmed our findings in RA-SH-SY5Y cells, but hPCN were less sensitive to Aβ; therefore, a 20 (Aβ₄₀) and 50 (Aβ₄₂) times higher concentration was needed for inducing uptake. The blocking of endocytosis completely inhibited the internalisation of Aβ peptides in RA-SH-SY5Y cells and hPCN, indicating that this is a major pathway by which Aβ enters the cells. In addition, the internalisation of Aβ₄₂, but not Aβ₄₀, was reduced by 55 % by blocking RAGE. Finally, for the first time we showed that pore formation in cell membranes by perforin led to Aβ internalisation in hPCN. Understanding how Aβ is internalised sheds light on the pathological role of Aβ and provides further ideas of inhibitory strategies for preventing Aβ internalisation and the spreading of neurodegeneration in AD.     

Long-Term GABA Administration Induces Alpha Cell-Mediated Beta-like Cell Neogenesis

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