Vitamin D deficiency is a worldwide pandemic. The aim of this study was to evaluate associations of plasma 25(OH)D levels with the likelihood of coronavirus disease 2019 (COVID‐19) infection and hospitalization. The study population included the 14 000 members of Leumit Health Services, who were tested for COVID‐19 infection from February 1st to April 30th, 2020, and who had at least one previous blood test for the plasma 25(OH)D level. ‘Suboptimal’ or ‘low’ plasma 25(OH)D level was defined as plasma 25‐hydroxyvitamin D, or 25(OH)D, concentration below the level of 30 ng/mL. Of 7807 individuals, 782 (10.02%) were COVID‐19‐positive, and 7025 (89.98%) COVID‐19‐negative. The mean plasma vitamin D level was significantly lower among those who tested positive than negative for COVID‐19 [19.00 ng/mL (95% confidence interval (CI) 18.41–19.59) vs. 20.55 (95% CI: 20.32–20.78)]. Univariate analysis demonstrated an association between the low plasma 25(OH)D level and increased likelihood of COVID‐19 infection [crude odds ratio (OR) of 1.58 (95% CI: 1.24–2.01, P < 0.001)], and of hospitalization due to the SARS‐CoV‐2 virus [crude OR of 2.09 (95% CI: 1.01–4.30, P < 0.05)]. In multivariate analyses that controlled for demographic variables, and psychiatric and somatic disorders, the adjusted OR of COVID‐19 infection [1.45 (95% CI: 1.08–1.95, P < 0.001)] and of hospitalization due to the SARS‐CoV‐2 virus [1.95 (95% CI: 0.98–4.845, P = 0.061)] were preserved. In the multivariate analyses, age over 50 years, male gender and low–medium socioeconomic status were also positively associated with the risk of COVID‐19 infection; age over 50 years was positively associated with the likelihood of hospitalization due to COVID‐19. We concluded that low plasma 25(OH)D levels appear to be an independent risk factor for COVID‐19 infection and hospitalization.

Solvent dynamics can play a major role in enzyme activity, but obtaining an accurate, quantitative picture of solvent activity during catalysis is quite challenging. Here, we combine terahertz spectroscopy and X-ray absorption analyses to measure changes in the coupled water-protein motions during peptide hydrolysis by a zinc-dependent human metalloprotease. These changes were tightly correlated with rearrangements at the active site during the formation of productive enzyme-substrate intermediates and were different from those in an enzyme–inhibitor complex. Molecular dynamics simulations showed a steep gradient of fast-to-slow coupled protein-water motions around the protein, active site and substrate. Our results show that water retardation occurs before formation of the functional Michaelis complex. We propose that the observed gradient of coupled protein-water motions may assist enzyme-substrate interactions through water-polarizing mechanisms that are remotely mediated by the catalytic metal ion and the enzyme active site.

Aminoacyl-tRNA synthetases (aaRSs) exert control over the faithful transfer of amino acids onto cognate tRNAs. Since chemical structures of various amino acids closely resemble each other, it is difficult to discriminate between them. Editing activity has been evolved by certain aaRSs to resolve the problem. In this study, we determined the crystal structures of complexes of T. thermophilus phenylalanyl-tRNA synthetase (PheRS) with L-tyrosine, p-chloro-phenylalanine, and a nonhydrolyzable tyrosyl-adenylate analog. The structures demonstrate plasticity of the synthetic site capable of binding substrates larger than phenylalanine and provide a structural basis for the proofreading mechanism. The editing site is localized at the B3/B4 interface, 35 A from the synthetic site. Glubeta334 plays a crucial role in the specific recognition of the Tyr moiety in the editing site. The tyrosyl-adenylate analog binds exclusively in the synthetic site. Both structural data and tyrosine-dependent ATP hydrolysis enhanced by tRNA(Phe) provide evidence for a preferential posttransfer editing pathway in the phenylalanine-specific system.

Understanding the crystallization of organic molecules is a long-standing challenge. Herein, a mechanistic study on the self-assembly of crystalline arrays in aqueous solution is presented. The crystalline arrays are assembled from perylene diimide (PDI) amphiphiles bearing a chiral N-acetyltyrosine side group connected to the PDI aromatic core. A kinetic study of the crystallization process was performed using circular dichroism spectroscopy combined with time-resolved cryogenic transmission electron microscopy (cryo-TEM) imaging of key points along the reaction coordinate, and molecular dynamics simulation of the initial stages of the assembly. The study reveals a complex self-assembly process starting from the formation of amorphous aggregates that are transformed into crystalline material through a nucleation-growth process. Activation parameters indicate the key role of desolvation along the assembly pathway. The insights from the kinetic study correlate well with the structural data from cryo-TEM imaging. Overall, the study reveals four stages of crystalline self-assembly: 1) collapse into amorphous aggregates; 2) nucleation as partial ordering; 3) crystal growth; and 4) fusion of smaller crystalline aggregates into large crystals. These studies indicate that the assembly process proceeds according to a two-step crystallization model, whereby initially formed amorphous material is reorganized into an ordered system. This process follows Ostwald's rule of stages, evolving through a series of intermediate phases prior to forming the final structure, thus providing an insight into the crystalline self-assembly process in aqueous medium.

The crystal structure of the ternary complex of (alphabeta)(2) heterotetrameric phenylalanyl-tRNA synthetase (PheRS) from Thermus thermophilus with cognate tRNA(Phe) and a nonhydrolyzable phenylalanyl-adenylate analogue (PheOH-AMP) has been determined at 3.1 A resolution. It reveals conformational changes in tRNA(Phe) induced by the PheOH-AMP binding. The single-stranded 3' end exhibits a hairpin conformation in contrast to the partial unwinding observed previously in the binary PheRS.tRNA(Phe) complex. The CCA end orientation is stabilized by extensive base-specific interactions of A76 and C75 with the protein and by intra-RNA interactions of A73 with adjacent nucleotides. The 4-amino group of the "bulged out" C75 is trapped by two negatively charged residues of the beta subunit (Glubeta31 and Aspbeta33), highly conserved in eubacterial PheRSs. The position of the A76 base is stabilized by interactions with Hisalpha212 of motif 2 (universally conserved in PheRSs) and class II-invariant Argalpha321 of motif 3. Important conformational changes induced by the binding of tRNA(Phe) and PheOH-AMP are observed in the catalytic domain: the motif 2 loop and a "helical" loop (residues 139-152 of the alpha subunit) undergo coordinated displacement; Metalpha148 of the helical loop adopts a conformation preventing the 2'-OH group of A76 from approaching the alpha-carbonyl carbon of PheOH-AMP. The unfavorable position of the terminal ribose stems from the absence of the alpha-carbonyl oxygen in the analogue. Our data suggest that the idiosyncratic feature of PheRS, which aminoacylates the 2'-OH group of the terminal ribose, is dictated by the system-specific topology of the CCA end-binding site.

Protein flexibility is thought to play key roles in numerous biological processes, including antibody affinity maturation, signal transduction, and enzyme catalysis, yet only limited information is available regarding the molecular details linking protein dynamics with function. A single point mutation at the distal site of the endogenous tissue inhibitor of metalloproteinase 1 (TIMP-1) enables this clinical target protein to tightly bind and inhibit membrane type 1 matrix metalloproteinase (MT1-MMP) by increasing only the association constant. The high-resolution X-ray structure of this complex determined at 2 A could not explain the mechanism of enhanced binding and pointed to a role for protein conformational dynamics. Molecular dynamics (MD) simulations reveal that the high-affinity TIMP-1 mutants exhibit significantly reduced binding interface flexibility and more stable hydrogen bond networks. This was accompanied by a redistribution of the ensemble of substrates to favorable binding conformations that fit the enzyme catalytic site. Apparently, the decrease in backbone flexibility led to a lower entropy cost upon formation of the complex. This work quantifies the effect of a single point mutation on the protein conformational dynamics and function of TIMP-1. Here we argue that controlling the intrinsic protein dynamics of MMP endogenous inhibitors may be utilized for rationalizing the design of selective novel protein inhibitors for this class of enzymes.

No abstract

A new coronavirus infection, COVID-19, has recently emerged, and has caused a global pandemic along with an international public health emergency. Currently, no licensed vaccines are available for COVID-19. The identification of immunodominant epitopes for both B- and T-cells that induce protective responses in the host is crucial for effective vaccine design. Computational prediction of potential epitopes might significantly reduce the time required to screen peptide libraries as part of emergent vaccine design. In our present study, we used an extensive immunoinformatics-based approach to predict conserved immunodominant epitopes from the proteome of SARS-CoV-2. Regions from SARS-CoV-2 protein sequences were defined as immunodominant, based on the following three criteria regarding B- and T-cell epitopes: (i) they were both mapped, (ii) they predicted protective antigens, and (iii) they were completely identical to experimentally validated epitopes of SARS-CoV. Further, structural and molecular docking analyses were performed in order to understand the binding interactions of the identified immunodominant epitopes with human major histocompatibility complexes (MHC). Our study provides a set of potential immunodominant epitopes that could enable the generation of both antibody- and cell-mediated immunity. This could contribute to developing peptide vaccine-based adaptive immunotherapy against SARS-CoV-2 infections and prevent future pandemic outbreaks.