Metabolomics and NMR.

The purpose of this manuscript will be to convince the reader to dive deeper into NMR spectroscopy and prevent the technique from being just another "black-box" in the lab. We will try to concisely highlight interesting topics and supply additional references for further exploration at each stage. The advantages of delving into the technique will be shown. The secondary objective, i.e., avoiding common problems before starting, will hopefully then become clear. Lastly, we will emphasize the spectrometer information needed for manuscript reporting to allow reproduction of results and confirm findings.

NMR as a "Gold Standard" Method in Drug Design and Discovery.

Studying disease models at the molecular level is vital for drug development in order to improve treatment and prevent a wide range of human pathologies. Microbial infections are still a major challenge because pathogens rapidly and continually evolve developing drug resistance. Cancer cells also change genetically, and current therapeutic techniques may be (or may become) ineffective in many cases. The pathology of many neurological diseases remains an enigma, and the exact etiology and underlying mechanisms are still largely unknown. Viral infections spread and develop much more quickly than does the corresponding research needed to prevent and combat these infections; the present and most relevant outbreak of SARS-CoV-2, which originated in Wuhan, China, illustrates the critical and immediate need to improve drug design and development techniques. Modern day drug discovery is a time-consuming, expensive process. Each new drug takes in excess of 10 years to develop and costs on average more than a billion US dollars. This demonstrates the need of a complete redesign or novel strategies. Nuclear Magnetic Resonance (NMR) has played a critical role in drug discovery ever since its introduction several decades ago. In just three decades, NMR has become a "gold standard" platform technology in medical and pharmacology studies. In this review, we present the major applications of NMR spectroscopy in medical drug discovery and development. The basic concepts, theories, and applications of the most commonly used NMR techniques are presented. We also summarize the advantages and limitations of the primary NMR methods in drug development.

Looking for Mr(s) Right: Decision bias can prevent us from finding the most attractive face.

In realistic and challenging decision contexts, people may show biases that prevent them from choosing their favored options. For example, astronomer Johannes Kepler famously interviewed several candidate fiancées sequentially, but was rejected when attempting to return to a previous candidate. Similarly, we examined human performance on searches for attractive faces through fixed-length sequences by adapting optimal stopping computational theory developed from behavioral ecology and economics. Although economics studies have repeatedly found that participants sample too few options before choosing the best-ranked number from a series, we instead found overlong searches with many sequences ending without choice. Participants employed irrationally high choice thresholds, compared to the more lax, realistic standards of a Bayesian ideal observer, which achieved better-ranked faces. We consider several computational accounts and find that participants most resemble a Bayesian model that decides based on altered attractiveness values. These values may produce starkly different biases in the facial attractiveness domain than in other decision domains.

Long TE STEAM and PRESS for estimating fat olefinic/methyl ratios and relative ω-3 fat content at 3T.

BACKGROUND: Fat olefinic/methyl ratios provide a measure of fat unsaturation. The methyl resonance linewidth is altered with the presence of ω-3 fat. PURPOSE: To optimize stimulated echo acquisition mode (STEAM) and point resolved spectroscopy (PRESS) echo times (TE) at 3T to 1) improve olefinic/methyl ratios and 2) enable relative ω-3 fat content assessment. STUDY TYPE: Technical development on phantoms and healthy volunteers. POPULATION: Nine edible oils and four healthy volunteers (tibial bone marrow). FIELD STRENGTH/SEQUENCE: STEAM (mixing time = 20 msec) and PRESS sequences at 3T. High-resolution oil spectra at 16.5T. ASSESSMENT: 3T STEAM and PRESS olefinic/methyl ratios as a function of TE were compared to 16.5T measures for the oils, and to a literature-deduced value for tibial bone marrow. Oil methyl linewidths were calculated at each TE to investigate correlation with expected ω-3 fatty acid content. STATISTICAL TESTS: Percent differences were calculated between oil olefinic/methyl ratios obtained at 3T and 16.5T. Linear regression R2 values measured correlation of methyl linewidth to ω-3 content. RESULTS: STEAM, TE = 120 msec, resulted in average oil olefinic/methyl ratios that differ by about -4.8% compared to high-resolution ratios. Tibial bone marrow olefinic/methyl ratios differ by -1.8% compared with literature-obtained ratios. PRESS, TE = 180 msec, resulted in oil ratios that differ by 7.8% and tibial bone marrow ratios that differ by 0.2%. A TE of 160 msec for both STEAM and PRESS enabled relative levels of oil ω-3 fatty acid content to be estimated (R2 values ≥0.9). DATA CONCLUSION: STEAM, TE = 120 msec (mixing time = 20 msec), and PRESS, TE = 180 msec, optimally estimated olefinic/methyl ratios. STEAM and PRESS, TE = 160 msec, enable relative oil ω-3 fatty acid estimation from methyl linewidths. LEVEL OF EVIDENCE: 1 Technical Efficacy Stage 1 J. Magn. Reson. Imaging 2017.

Do the folk actually hold folk-economic beliefs?

Boyer & Petersen (B&P) argue that folk-economic beliefs are widespread - shaped by evolved cognitive systems - and they offer exemplar beliefs to illustrate their thesis. In this commentary, we highlight evidence of substantial variation in one of these exemplars: beliefs about immigration. Contra claims by B&P, we argue that the balance of this evidence suggests the "folk" may actually hold positive beliefs about the economic impact of immigration.

Insights into the Mechanism of Action of the Two-Peptide Lantibiotic Lacticin 3147.

Lacticin 3147 is a two peptide lantibiotc (LtnA1 and LtnA2) that displays nanomolar activity against many Gram-positive bacteria. Lacticin 3147 may exert its antimicrobial effect by several mechanisms. Isothermal titration calorimetry experiments show that only LtnA1 binds to the peptidoglycan precursor lipid II, which could inhibit peptidoglycan biosynthesis. An experimentally supported model of the resulting complex suggests that the key binding partners are the C-terminus of LtnA1 and pyrophosphate of lipid II. A combination of in vivo and in vitro assays indicates that LtnA1 and LtnA2 can induce rapid membrane lysis without the need for lipid II binding. However, the presence of lipid II substantially increases the activity of lacticin 3147. Furthermore, studies with synthetic LtnA2 analogues containing either desmethyl- or oxa-lanthionine rings confirm that the precise geometry of these rings is essential for this synergistic activity.

Choosing the right level of analysis: Stereotypes shape social reality via collective action.

In his 2012 book Jussim argues that the self-fulfilling prophecy and expectancy effects of descriptive stereotypes are not potent shapers of social reality. However, his conclusion that descriptive stereotypes per se do not shape social reality is premature and overly reductionist. We review evidence that suggests descriptive stereotypes do have a substantial influence on social reality, by virtue of their influence on collective action.

Recommendations and Standardization of Biomarker Quantification Using NMR-Based Metabolomics with Particular Focus on Urinary Analysis.

NMR-based metabolomics has shown considerable promise in disease diagnosis and biomarker discovery because it allows one to nondestructively identify and quantify large numbers of novel metabolite biomarkers in both biofluids and tissues. Precise metabolite quantification is a prerequisite to move any chemical biomarker or biomarker panel from the lab to the clinic. Among the biofluids commonly used for disease diagnosis and prognosis, urine has several advantages. It is abundant, sterile, and easily obtained, needs little sample preparation, and does not require invasive medical procedures for collection. Furthermore, urine captures and concentrates many "unwanted" or "undesirable" compounds throughout the body, providing a rich source of potentially useful disease biomarkers; however, incredible variation in urine chemical concentrations makes analysis of urine and identification of useful urinary biomarkers by NMR challenging. We discuss a number of the most significant issues regarding NMR-based urinary metabolomics with specific emphasis on metabolite quantification for disease biomarker applications and propose data collection and instrumental recommendations regarding NMR pulse sequences, acceptable acquisition parameter ranges, relaxation effects on quantitation, proper handling of instrumental differences, sample preparation, and biomarker assessment.

Solution structure of acidocin B, a circular bacteriocin produced by Lactobacillus acidophilus M46.

Acidocin B, a bacteriocin produced by Lactobacillus acidophilus M46, was originally reported to be a linear peptide composed of 59 amino acid residues. However, its high sequence similarity to gassericin A, a circular bacteriocin from Lactobacillus gasseri LA39, suggested that acidocin B might be circular as well. Acidocin B was purified from culture supernatant by a series of hydrophobic interaction chromatographic steps. Its circular nature was ascertained by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry and tandem mass spectrometry (MS/MS) sequencing. The peptide sequence was found to consist of 58 amino acids with a molecular mass of 5,621.5 Da. The sequence of the acidocin B biosynthetic gene cluster was also determined and showed high nucleotide sequence similarity to that of gassericin A. The nuclear magnetic resonance (NMR) solution structure of acidocin B in sodium dodecyl sulfate micelles was elucidated, revealing that it is composed of four α-helices of similar length that are folded to form a compact, globular bundle with a central pore. This is a three-dimensional structure for a member of subgroup II circular bacteriocins, which are classified based on their isoelectric points of ∼7 or lower. Comparison of acidocin B with carnocyclin A, a subgroup I circular bacteriocin with four α-helices and a pI of 10, revealed differences in the overall folding. The observed variations could be attributed to inherent diversity in their physical properties, which also required the use of different solvent systems for three-dimensional structural elucidation.

Paranoid males have reduced lateralisation for processing of negative emotions: an investigation using the chimeric faces test.

Reduced strength of lateralisation in patients with schizophrenia has been reported in a number of studies. However the exact nature of this relationship remains unclear. In this study, lateralisation for processing emotional faces was measured using the chimeric faces test and examined in relation to paranoia in a non-clinical sample. For males only, those with higher scores on a paranoia questionnaire had reduced lateralisation for processing negative facial emotion. For females there were no significant relationships. These findings suggest that atypical patterns of lateralisation for processing emotional stimuli may be implicated in, or associated with, increased levels of paranoia.

Solution structures of the linear leaderless bacteriocins enterocin 7A and 7B resemble carnocyclin A, a circular antimicrobial peptide.

Leaderless bacteriocins are a class of ribosomally synthesized antimicrobial peptides that are produced by certain Gram-positive bacteria without an N-terminal leader section. These bacteriocins are of great interest due to their potent inhibition of many Gram-positive organisms, including food-borne pathogens such as Listeria and Clostridium spp. We now report the NMR solution structures of enterocins 7A and 7B, leaderless bacteriocins recently isolated from Enterococcus faecalis 710C. These are the first three-dimensional structures to be reported for bacteriocins of this class. Unlike most other linear Gram-positive bacteriocins, enterocins 7A and 7B are highly structured in aqueous conditions. Both peptides are primarily α-helical, adopting a similar overall fold. The structures can be divided into three separate α-helical regions: the N- and C-termini are both α-helical, separated by a central kinked α-helix. The overall structures bear an unexpected resemblance to carnocyclin A, a 60-residue peptide that is cyclized via an amide bond between the C- and N-termini and has a saposin fold. Because of synergism observed for other two-peptide leaderless bacteriocins, it was of interest to probe possible binding interactions between enterocins 7A and 7B. However, despite synergistic activity observed between these peptides, no significant binding interaction was observed based on NMR and isothermal calorimetry.

'So Help Me God'? Does oath swearing in courtroom scenarios impact trial outcomes?

In countries such as Britain and the US, court witnesses must declare they will provide truthful evidence and are often compelled to publicly choose between religious ("oath") and secular ("affirmation") versions of this declaration. Might defendants who opt to swear an oath enjoy more favourable outcomes than those who choose to affirm? Two preliminary, pre-registered survey studies using minimal vignettes (Study 1, N = 443; Study 2, N = 913) indicated that people associate choice of the oath with credible testimony; and that participants, especially religious participants, discriminate against defendants who affirm. In a third, Registered Report study (Study 3, N = 1821), we used a more elaborate audiovisual mock trial paradigm to better estimate the real-world influence of declaration choice. Participants were asked to render a verdict for a defendant who either swore or affirmed, and were themselves required to swear or affirm that they would try the defendant in good faith. Overall, the defendant was not considered guiltier when affirming rather than swearing, nor did mock-juror belief in God moderate this effect. However, jurors who themselves swore an oath did discriminate against the affirming defendant. Exploratory analyses suggest this effect may be driven by authoritarianism, perhaps because high-authoritarian jurors consider the oath the traditional (and therefore correct) declaration to choose. We discuss the real-world implications of these findings and conclude the religious oath is an antiquated legal ritual that needs reform.

COVID-19 and seasonal flu vaccination hesitancy: Links to personality and general intelligence in a large, UK cohort.

Vaccines are a powerful and relatively safe tool to protect against a range of serious diseases. Nonetheless, a sizeable minority of people express 'vaccination hesitancy'. Accordingly, understanding the bases of this hesitancy represents a significant public health opportunity. In the present study we sought to examine the role of Big Five personality traits and general intelligence as predictors of vaccination hesitancy across two vaccination types in a large (N = 9667) sample of UK adults drawn from the Understanding Society longitudinal household study. We found that lower levels of general intelligence were associated with COVID-19 and seasonal flu vaccination hesitancy, and lower levels of neuroticism was associated with COVID-19 vaccination hesitancy. Although the self-reported reasons for being vaccine hesitant indicated a range of factors were important to people, lower general intelligence was associated with virtually all of these reasons. In contrast, Big Five personality traits showed more nuanced patterns of association.

The 3D structure of thuricin CD, a two-component bacteriocin with cysteine sulfur to α-carbon cross-links.

Thuricin CD is an antimicrobial factor that consists of two peptides, Trn-α and Trn-ß, that exhibit synergistic activity against drug resistant strains of Clostridium difficile. Trn-α and Trn-ß each possess three sulfur to α-carbon thioether bridges for which the stereochemistry is unknown. This report presents the three-dimensional solution structures of Trn-α and Trn-ß. Structure calculations were performed for the eight possible stereoisomers of each peptide based on the same NMR data. The structure of the stereoisomer that best fit the experimental data was chosen as the representative structure for each peptide. It was determined that Trn-α has L-stereochemistry at Ser21 (α-R), L-stereochemistry at Thr25 (α-R), and D-stereochemistry at Thr28 (α-S) (an LLD isomer). Trn-ß was also found to be the LLD isomer, with L-stereochemistry at Thr21 (α-R), L-stereochemistry at Ala25 (α-R), and D-stereochemistry at Tyr28 (α-S).

Religion and delusion.

We review scholarship that examines relationships - and distinctions - between religion and delusion. We begin by outlining and endorsing the position that both involve belief. Next, we present the prevailing psychiatric view that religious beliefs are not delusional if they are culturally accepted. While this cultural exemption has controversial implications, we argue it is clinically valuable and consistent with a growing awareness of the social - as opposed to purely epistemic - function of belief formation. Finally, we review research on continuities between religious and delusional cognition, which reveals that religious content is quite common in delusions and which provides tentative evidence for a positive relationship between religious belief and delusion-like belief in the general population.

N-Terminal Finger Stabilizes the S1 Pocket for the Reversible Feline Drug GC376 in the SARS-CoV-2 Mpro Dimer.

The main protease (Mpro, also known as 3CL protease) of SARS-CoV-2 is a high priority drug target in the development of antivirals to combat COVID-19 infections. A feline coronavirus antiviral drug, GC376, has been shown to be effective in inhibiting the SARS-CoV-2 main protease and live virus growth. As this drug moves into clinical trials, further characterization of GC376 with the main protease of coronaviruses is required to gain insight into the drug's properties, such as reversibility and broad specificity. Reversibility is an important factor for therapeutic proteolytic inhibitors to prevent toxicity due to off-target effects. Here we demonstrate that GC376 has nanomolar Ki values with the Mpro from both SARS-CoV-2 and SARS-CoV strains. Restoring enzymatic activity after inhibition by GC376 demonstrates reversible binding with both proteases. In addition, the stability and thermodynamic parameters of both proteases were studied to shed light on physical chemical properties of these viral enzymes, revealing higher stability for SARS-CoV-2 Mpro. The comparison of a new X-ray crystal structure of Mpro from SARS-CoV complexed with GC376 reveals similar molecular mechanism of inhibition compared to SARS-CoV-2 Mpro, and gives insight into the broad specificity properties of this drug. In both structures, we observe domain swapping of the N-termini in the dimer of the Mpro, which facilitates coordination of the drug's P1 position. These results validate that GC376 is a drug with an off-rate suitable for clinical trials.

Catalytic proficiency of ubiquitin conjugation enzymes: balancing pK(a) suppression, entropy, and electrostatics.

Biological organisms orchestrate coordinated responses to external stimuli through temporal fluctuations in protein-protein interaction networks using molecular mechanisms such as the synthesis and recognition of polyubiquitin (polyUb) chains on signaling adaptor proteins. One of the pivotal chemical steps in ubiquitination involves reaction of a lysine amino group with a thioester group on an activated E2, or ubiquitin conjugation enzyme, to form an amide bond between Ub and a target protein. In this study, we demonstrate a nominal 14-fold range for the rate of the chemical step, k(cat), catalyzed by different E2 enzymes using non-steady-state, single-turnover assays. However, the observed range for k(cat) is as large as ∼100-fold for steady-state, single-turnover assays. Biochemical assays were used in combination with measurement of the underlying protein-protein interaction kinetics using NMR line-shape and ZZ-exchange analyses to determine the rate of polyUb chain synthesis catalyzed by the heterodimeric E2 enzyme Ubc13-Mms2. Modest variations in substrate affinity and k(cat) can achieve functional diversity in E2 mechanism, thereby influencing the biological outcomes of polyubiquitination. E2 enzymes achieve reaction rate enhancements through electrostatic effects such as suppression of substrate lysine pK(a) and stabilization of transition states by the preorganized, polar enzyme active site as well as the entropic effects of binding. Importantly, modestly proficient enzymes such as E2s maintain the ability to tune reaction rates; this may confer a biological advantage for achieving specificity in the diverse cellular roles for which these enzymes are involved.

Feline coronavirus drug inhibits the main protease of SARS-CoV-2 and blocks virus replication.

The main protease, Mpro (or 3CLpro) in SARS-CoV-2 is a viable drug target because of its essential role in the cleavage of the virus polypeptide. Feline infectious peritonitis, a fatal coronavirus infection in cats, was successfully treated previously with a prodrug GC376, a dipeptide-based protease inhibitor. Here, we show the prodrug and its parent GC373, are effective inhibitors of the Mpro from both SARS-CoV and SARS-CoV-2 with IC50 values in the nanomolar range. Crystal structures of SARS-CoV-2 Mpro with these inhibitors have a covalent modification of the nucleophilic Cys145. NMR analysis reveals that inhibition proceeds via reversible formation of a hemithioacetal. GC373 and GC376 are potent inhibitors of SARS-CoV-2 replication in cell culture. They are strong drug candidates for the treatment of human coronavirus infections because they have already been successful in animals. The work here lays the framework for their use in human trials for the treatment of COVID-19.

Author Correction: Feline coronavirus drug inhibits the main protease of SARS-CoV-2 and blocks virus replication.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

The evolution of misbelief.

From an evolutionary standpoint, a default presumption is that true beliefs are adaptive and misbeliefs maladaptive. But if humans are biologically engineered to appraise the world accurately and to form true beliefs, how are we to explain the routine exceptions to this rule? How can we account for mistaken beliefs, bizarre delusions, and instances of self-deception? We explore this question in some detail. We begin by articulating a distinction between two general types of misbelief: those resulting from a breakdown in the normal functioning of the belief formation system (e.g., delusions) and those arising in the normal course of that system's operations (e.g., beliefs based on incomplete or inaccurate information). The former are instances of biological dysfunction or pathology, reflecting "culpable" limitations of evolutionary design. Although the latter category includes undesirable (but tolerable) by-products of "forgivably" limited design, our quarry is a contentious subclass of this category: misbeliefs best conceived as design features. Such misbeliefs, unlike occasional lucky falsehoods, would have been systematically adaptive in the evolutionary past. Such misbeliefs, furthermore, would not be reducible to judicious - but doxastically noncommittal - action policies. Finally, such misbeliefs would have been adaptive in themselves, constituting more than mere by-products of adaptively biased misbelief-producing systems. We explore a range of potential candidates for evolved misbelief, and conclude that, of those surveyed, only positive illusions meet our criteria.