Use of an Improved Matching Algorithm to Select Scaffolds for Enzyme Design Based on a Complex Active Site Model

<div><p>Active site preorganization helps native enzymes electrostatically stabilize the transition state better than the ground state for their primary substrates and achieve significant rate enhancement. In this report, we hypothesize that a complex active site model for active site preorganization modeling should help to create preorganized active site design and afford higher starting activities towards target reactions. Our matching algorithm ProdaMatch was improved by invoking effective pruning strategies and the native active sites for ten scaffolds in a benchmark test set were reproduced. The root-mean squared deviations between the matched transition states and those in the crystal structures were < 1.0 Å for the ten scaffolds, and the repacking calculation results showed that 91% of the hydrogen bonds within the active sites are recovered, indicating that the active sites can be preorganized based on the predicted positions of transition states. The application of the complex active site model for <i>de novo</i> enzyme design was evaluated by scaffold selection using a classic catalytic triad motif for the hydrolysis of <i>p</i>-nitrophenyl acetate. Eighty scaffolds were identified from a scaffold library with 1,491 proteins and four scaffolds were native esterase. Furthermore, enzyme design for complicated substrates was investigated for the hydrolysis of cephalexin using scaffold selection based on two different catalytic motifs. Only three scaffolds were identified from the scaffold library by virtue of the classic catalytic triad-based motif. In contrast, 40 scaffolds were identified using a more flexible, but still preorganized catalytic motif, where one scaffold corresponded to the α-amino acid ester hydrolase that catalyzes the hydrolysis and synthesis of cephalexin. Thus, the complex active site modeling approach for <i>de novo</i> enzyme design with the aid of the improved ProdaMatch program is a promising approach for the creation of active sites with high catalytic efficiencies towards target reactions.</p></div

Side chain repacking results for seven scaffolds.

<p>Side chain repacking results for seven scaffolds.</p

U-shaped association between 25-hydroxyvitamin D concentration and the prevalence of asthma in the overweight and obese U.S. population

Vitamin D supplementation has shown promise in averting asthma. However, the association between 25(OH)D levels and asthma prevention in various demographic groups remains inconclusive. Here, we explore this relationship in the context of overweight and obese individuals in the United States. We scrutinized cross-sectional data derived from the National Health and Nutrition Examination Survey conducted between 2007 and 2018. This dataset encompasses comprehensive information about asthma patients with a body mass index greater than 25 kg/m2, in addition to data regarding 25(OH)D concentration and other pertinent variables. Among the 3889 participants, 16.2% (631/3889) reported a history of asthma, constituting 1765 (45.4%) males and 2124 (54.6%) females. The median age was 56.0 years, with a standard deviation of 16.0 years. We conducted restricted cubic spline (RCS) regression analysis to assess the correlation between 25(OH)D levels and asthma. After adjusting for confounders, compared to individuals with lower 25-hydroxyvitamin D concentration (group1 ≤ 46.6 nmol/L), the adjusted odds ratios (OR) for asthma in group2 (46.7–62 nmol/L), group3 (62.1–78.2 nmol/L), and group4 (≥78.3 nmol/L) were 0.68 (95% CI: 0.49–0.94, p p p p = 0.017) with an inflection point at approximately 84.95 nmol/L. High levels of 25(OH)D are correlated with a diminished prevalence of asthma among overweight and obese individuals in the United States. Vitamin D supplementation may reduce the incidence of asthma in the overweight and obese individuals.A u-shaped association was observed between 25-hydroxyvitamin D levels and asthma, with an inflection point of 84.95 nmol/L.Presenting supportive evidence for vitamin D supplementation in the overweight and obese individuals. Vitamin D supplementation may reduce the incidence of asthma in the overweight and obese individuals. A u-shaped association was observed between 25-hydroxyvitamin D levels and asthma, with an inflection point of 84.95 nmol/L. Presenting supportive evidence for vitamin D supplementation in the overweight and obese individuals.</p

Matching results and active site residue sidechain repacking results in scaffold 1mpx.

<p>(A) Superposition of native and matched active sites for hydrolysis of cephalexin on scaffold 1mpx. (B) Conformations of repacked residues based on matched cephalexin on scaffold 1mpx. The transition states are shown in ball and stick model and colored in pink. The active site residues are shown in stick model. Atoms O, N, and C in crystal structures are colored in red, teal, and gray, respectively. Matched residues are colored in red. The hydrogen bonds in crystal structures are shown in dotted green lines, and the predicted hydrogen bonds are shown in dotted pink lines. The distances between hydrogen bonding donors and acceptors are shown in Å and labeled besides the dotted lines.</p

Use of an Improved Matching Algorithm to Select Scaffolds for Enzyme Design Based on a Complex Active Site Model - Fig 3

<p><b>Side chain repacking results for scaffolds: (A) 1c2t; (B) 1jcl.</b> The transition states are shown in ball and stick model, and the active site residues in stick model. Atoms O, N, and C in crystal structures are colored in red, teal, and gray, respectively. The matched TS and repacked residues are colored in orange. The hydrogen bonds in crystal structures are shown in dotted green lines, and the predicted hydrogen bonds are shown in dotted orange lines. The distances between hydrogen bonding donors and acceptors are shown in Å and labeled besides the dotted lines.</p

Selected scaffolds by ProdaMatch for hydrolytic reactions of PNPA and cephalexin using catalytic triad based motifs.

<p>Selected scaffolds by ProdaMatch for hydrolytic reactions of PNPA and cephalexin using catalytic triad based motifs.</p

Superposition of native and predicted active sites.

<p><b>(A) 1c2t; (B) 1jcl.</b> The transition states are shown in ball and stick model, and the active site residues in stick model. Atoms O, N, and C in crystal structures are colored in red, teal, and gray, respectively. The matched structures based on complex active site model are shown in pink, and in green for matched structures based on minimal active site model.</p

Complex active site models for PNPA and cephalexin based on different catalytic motifs.

<p>(A) Classic catalytic triad motif for hydrolysis of PNPA; (B) Classic catalytic triad motif for hydrolysis of cephalexin; (C) Flexible catalytic triad motif for hydrolysis of cephalexin.</p

Recapitulation of native active sites by ProdaMatch for ten scaffolds.

<p>Recapitulation of native active sites by ProdaMatch for ten scaffolds.</p

Data Availability from How silanization influences aggregation and moisture sorption behaviours of silanized silica: analysis of porosity and multilayer moisture adsorption

Based on the results of nitrogen adsorption and dynamic vapour sorption as well as analysis by the Hailwood–Horrobin (H-H) model, the effects of γ-methacryloxypropyltrimethoxysilane (MPTS) on the agglomeration and moisture sorption properties of fumed silica particles were investigated. After adding various concentrations (2%, 4%, 6% and 8%) of MPTS, different degrees of silanization were obtained by showing various ─OH group contents on the silica surface, which resulted in silica agglomerates with different porous structures. The bigger mesopores in the unmodified silica agglomerates became smaller and finally disappeared after MPTS modification and the Bruanuer–Emmett–Teller surface area decreased more gradually with an increase in MPTS concentration. The H-H model fitted the sorption isotherms very well, and both hydrated water and dissolved water showed decreasing trends with the increase in MPTS concentration, showing reduced hygroscopicity. Up to 6% MPTS, the ─OH groups decreased with increasing MPTS concentration, as indicated by reduced <i>K</i>_h and <i>W</i> parameters, while at 8% MPTS an extensive self-condensation of MPTS occurred. Adsorption hysteresis appeared for moisture sorption on silanized silica, especially at low relative humidity values and at low MPTS concentrations, which could be explained by a synergistic effect of the surface ─OH group content and pore characteristics. These results could aid our understanding of the applications of silane-modified silica particles