Trichome patterning in Arabidopsis is regulated by R2R3MYB, bHLH and WDR (MBW) genes. These are considered to form a trimeric MBW protein complex that promotes trichome formation. The MBW proteins are engaged in a regulatory network to select trichome cells among epidermal cells through R3MYB proteins that can move between cells and repress the MBW complex by competitive binding with the R2R3MYB to the bHLHL protein. We use quantitative pull-down assays to determine the relative dissociation constants for the protein-protein interactions of the involved genes. We find similar binding strength between the trichome promoting genes and weaker binding of the R3MYB inhibitors. We used the dissociation constants to calculate the relative percentage of all possible complex combinations and found surprisingly low fractions of those complexes that are typically considered to be relevant for the regulation events. Finally, we predict an increased robustness in patterning as a consequence of higher ordered complexes mediated by GL3 dimerization.

The interaction between SARS-CoV-2 spike RBD and ACE2 proteins is a crucial step for host cell infection by the virus. Without it, the entire virion entrance mechanism is compromised. The aim of this study was to evaluate the capacity of various natural product classes, including flavonoids, anthraquinones, saponins, ivermectin, chloroquine, and erythromycin, to modulate this interaction. To accomplish this, we applied a recently developed a microfluidic diffusional sizing (MDS) technique that allows us to probe protein-protein interactions via measurements of the hydrodynamic radius (Rh) and dissociation constant (KD); the evolution of Rh is monitored in the presence of increasing concentrations of the partner protein (ACE2); and the KD is determined through a binding curve experimental design. In a second time, with the protein partners present in equimolar amounts, the Rh of the protein complex was measured in the presence of different natural products. Five of the nine natural products/extracts tested were found to modulate the formation of the protein complex. A methanol extract of Chenopodium quinoa Willd bitter seed husks (50 µg/mL; bisdesmoside saponins) and the flavonoid naringenin (1 µM) were particularly effective. This rapid selection of effective modulators will allow us to better understand agents that may prevent SARS-CoV-2 infection.

Electrochemical sensors that incorporate immunoassay principles have the ability to monitor dynamic processes of antigen-antibody interactions in real time. In this study, a gold electrode was modified with tin dioxide colloidal quantum wire (SnO2 QWs) and then coated with the leucine/arginine subtype microcystin (MC-LR) antibody. The active site of SnO2 QWs that was not bound by MC-LR antibody was then passivated with bovine serum protein (BSA). When the MC-LR antigen binds specifically to the antibodies on the electrode\'s surface, it triggers electrochemical reactions and generates electrical signals at specific voltage conditions. The SnO2 QW exhibits excellent electron transport ability, and its ability to form a loose and porous microstructure on the gold electrode surface, which is conducive to the receptor function of the biosensor. The results show a high affinity between the MC-LR antigen and antibody, ranging from 1 pg/mL to 10 ng/mL of MC-LR antigen concentration. The kinetic characteristics of the immune reaction between MC-LR antigen and antibody were elucidated, obtaining a binding constant of 1.399 × 1011 M-1 and a dissociation constant of 7.147 pM, demonstrating the potential of electrochemical biosensing technology in biomolecular interactions.

BACKGROUND: Amino acid property-aware phylogenetic analysis (APPA) refers to the phylogenetic analysis method based on amino acid property encoding, which is used for understanding and inferring evolutionary relationships between species from the molecular perspective. Fast Fourier transform (FFT) and Higuchi\'s fractal dimension (HFD) have excellent performance in describing sequences\' structural and complexity information for APPA. However, with the exponential growth of protein sequence data, it is very important to develop a reliable APPA method for protein sequence analysis.
RESULTS: Consequently, we propose a new method named FFP, it joints FFT and HFD. Firstly, FFP is used to encode protein sequences on the basis of the important physicochemical properties of amino acids, the dissociation constant, which determines acidity and basicity of protein molecules. Secondly, FFT and HFD are used to generate the feature vectors of encoded sequences, whereafter, the distance matrix is calculated from the cosine function, which describes the degree of similarity between species. The smaller the distance between them, the more similar they are. Finally, the phylogenetic tree is constructed. When FFP is tested for phylogenetic analysis on four groups of protein sequences, the results are obviously better than other comparisons, with the highest accuracy up to more than 97%.
CONCLUSIONS: FFP has higher accuracy in APPA and multi-sequence alignment. It also can measure the protein sequence similarity effectively. And it is hoped to play a role in APPA\'s related research.

With the emergence of Delta and Omicron variants, many other important variants of SARS-CoV-2, which cause Coronavirus disease-2019, including A.30, are reported to increase the concern created by the global pandemic. The A.30 variant, reported in Tanzania and other countries, harbors spike gene mutations that help this strain to bind more robustly and to escape neutralizing antibodies. The present study uses molecular modelling and simulation-based approaches to investigate the key features of this strain that result in greater infectivity. The protein-protein docking results for the spike protein demonstrated that additional interactions, particularly two salt-bridges formed by the mutated residue Lys484, increase binding affinity, while the loss of key residues at the N terminal domain (NTD) result in a change to binding conformation with monoclonal antibodies, thus escaping their neutralizing effects. Moreover, we deeply studied the atomic features of these binding complexes through molecular simulation, which revealed differential dynamics when compared to wild type. Analysis of the binding free energy using MM/GBSA revealed that the total binding free energy (TBE) for the wild type receptor-binding domain (RBD) complex was -58.25 kcal/mol in contrast to the A.30 RBD complex, which reported -65.59 kcal/mol. The higher TBE for the A.30 RBD complex signifies a more robust interaction between A.30 variant RBD with ACE2 than the wild type, allowing the variant to bind and spread more promptly. The BFE for the wild type NTD complex was calculated to be -65.76 kcal/mol, while the A.30 NTD complex was estimated to be -49.35 kcal/mol. This shows the impact of the reported substitutions and deletions in the NTD of A.30 variant, which consequently reduce the binding of mAb, allowing it to evade the immune response of the host. The reported results will aid the development of cross-protective drugs against SARS-CoV-2 and its variants.

Smoothened (SMO) protein is a member of the G protein-coupled receptor (GPCR) family that is involved in the Hedgehog (Hh) signaling pathway. It is a putative target for treating various cancers, including medulloblastoma and basal cell carcinoma (BCC). Characterizing membrane proteins such as SMO in their native state is highly beneficial for the development of effective pharmaceutical drugs, as their structures and functions are retained to the highest extent in this state. Therefore, although SMO protein is conventionally solubilized in detergent micelles, incorporating the protein in a lipid-based membrane mimic is still required. In this study, we used styrene maleic acid (SMA) copolymer that directly extracted membrane protein and surrounding lipids as well as formed the so-called polymer nanodiscs, to solubilize and purify the SMO transmembrane domain encapsulated by SMA-nanodiscs. The obtained SMA-nanodiscs showed high homogeneity and maintained the physiological activity of SMO protein, thereby enabling the measurement of the dissociation constant (Kd) for SMO ligands SMO-ligands Shh Signaling Antagonist V (SANT-1) and Smoothened Agonist (SAG) using ligand-based solution nuclear magnetic resonance spectroscopy. This work paves the way for investigating the structure, function, and drug development of SMO proteins in a native-like lipid environment.

Saturation transfer MRI has previously been used to probe molecular binding interactions with signal enhancement via the water signal. Here, we detail the relayed nuclear overhauser effect (rNOE) based mechanisms of this signal enhancement, develop a strategy of quantifying molecular binding affinity, i.e., the dissociation constant ( KD$$ {K}_D $$ ), and apply the method to detect electrostatic binding of several charged small biomolecules. Another goal was to estimate the detection limit for transient receptor-substrate binding.
The signal enhancement mechanism was quantitatively described by a three-step magnetization transfer model, and numerical simulations were performed to verify this theory. The binding equilibria of arginine, choline, and acetyl-choline to anionic resin were studied as a function of ligand concentration, pH, and salt content. Equilibrium dissociation constants ( KD$$ {K}_D $$ ) were determined by fitting the multiple concentration data.
The numerical simulations indicate that the signal enhancement is sufficient to detect the molecular binding of sub-millimolar (∼100 μM) concentration ligands to low micromolar levels of molecular targets. The measured rNOE signals from arginine, choline, and acetyl-choline binding experiments show that several magnetization transfer pathways (intra-ligand rNOEs and intermolecular rNOEs) can contribute. The rNOEs that arise from molecular ionic binding were influenced by pH and salt concentration. The molecular binding strengths in terms of KD$$ {K}_{\\mathrm{D}} $$ ranged from 70-160 mM for the three cations studied.
The capability to use MRI to detect the transient binding of small substrates paves a pathway towards the detection of micromolar level receptor-substrate binding in vivo.

As SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) continues to inflict chaos globally, a new variant officially known as B.1.1.529 was reported in South Africa and was found to harbor 30 mutations in the spike protein. It is too early to speculate on transmission and hospitalizations. Hence, more analyses are required, particularly to connect the genomic patterns to the phenotypic attributes to reveal the binding differences and antibody response for this variant, which can then be used for therapeutic interventions. Given the urgency of the required analysis and data on the B.1.1.529 variant, we have performed a detailed investigation to provide an understanding of the impact of these novel mutations on the structure, function, and binding of RBD to hACE2 and mAb to the NTD of the spike protein. The differences in the binding pattern between the wild type and B.1.1.529 variant complexes revealed that the key substitutions Asn417, Ser446, Arg493, and Arg498 in the B.1.1.529 RBD caused additional interactions with hACE2 and the loss of key residues in the B.1.1.529 NTD resulted in decreased interactions with three CDR regions (1-3) in the mAb. Further investigation revealed that B.1.1.529 displayed a stable dynamic that follows a global stability trend. In addition, the dissociation constant (KD), hydrogen bonding analysis, and binding free energy calculations further validated the findings. Hydrogen bonding analysis demonstrated that significant hydrogen bonding reprogramming took place, which revealed key differences in the binding. The total binding free energy using MM/GBSA and MM/PBSA further validated the docking results and demonstrated significant variations in the binding. This study is the first to provide a basis for the higher infectivity of the new SARS-CoV-2 variants and provides a strong impetus for the development of novel drugs against them.

In capillary electrophoresis, determination of the basic physical and chemical properties of compounds, such as absolute mobility (m0) and dissociation constant (pKa), is of great practical significance. This is because the aforementioned properties are often used for the qualitative or quantitative analyses of the relevant compounds toward their application as potential drugs. Lovastatin is a potential drug candidate that can reduce the levels of cholesterol and low-density lipoprotein cholesterol in the blood, as well as prevent atherosclerosis and coronary heart disease. For a more convenient and rapid investigation of the properties and applications of lovastatin, it is necessary to determine its m0 and pKa values. However, existing research on capillary electrophoresis for lovastatin and other related drugs focus on their quantitative determination, and their action mechanism and functions. Unfortunately, there are very few studies aimed at the determination of the m0 and pKa values of lovastatin. Based on related studies, this paper herein proposed a novel method to determine m0 and pKa of lovastatin. The present study mainly included a calculation method and experimental verification. The calculation method was based on capillary zone electrophoresis (CZE) and the empirical formula of ion mobility. First, on the basis of the empirical formula, the calculation formula for m0 was derived from the relationship between the actual mobility (mact), effective mobility (meff) and m0. Second, for a monovalent acid (HA), according to the calculation formula for m0 part, considering the hydrogen ion concentration as the independent variable and the reciprocal of meff as the dependent variable, a straight line was obtained on the coordinate axis. From the slope of this straight line, the dissociation equilibrium constant Ka was obtained directly, and pKa was calculated easily. After the derivation of m0 and pKa in the theoretical part, the feasibility and reliability of this method were verified by using it to determine the m0 and pKa values of several organic acids and bases (barbituric acid, benzoic acid, benzylamine, phenol, and m-cresol) in the experimental part. Note that for the buffer system with pH<6.0, reverse capillary electrophoresis was used for the determination of pKa, because this technique helped shorten the migration time and facilitates the detection of analytes that could not reach the cathode. After obtaining m0 and pKa, the theoretical reference values for these parameters were obtained by PeakMaster 5.1. The experimental data were well consistent with the theoretical m0 and pKa values. The standard deviation (SDs) of m0 and pKa were less than 6.0% and 6.2%, respectively. From the correlation coefficient (R) of the linear regression equation, it was found that the linear regression lines of pKa fit well, indicating the excellent reliability of this method. Finally, with this simple and reliable method, dimethyl sulfoxide (DMSO) was used as a marker for electroosmotic flow to determine the m0 and pKa values of lovastatin (-1.70×10-8 m2/(V·s) and 9.00, respectively). This method is suitable for the determination of m0 and pKa of acidic and basic analytes. The method has high accuracy and is expected to play an indispensable role in drug analysis.

The evolution of new SARS-CoV-2 variants around the globe has made the COVID-19 pandemic more worrisome, further pressuring the health care system and immunity. Novel variations that are unique to the receptor-binding motif (RBM) of the receptor-binding domain (RBD) spike glycoprotein, i. e. L452R-E484Q, may play a different role in the B.1.617 (also known as G/452R.V3) variant\'s pathogenicity and better survival compared to the wild type. Therefore, a thorough analysis is needed to understand the impact of these mutations on binding with host receptor (RBD) and to guide new therapeutics development. In this study, we used structural and biomolecular simulation techniques to explore the impact of specific mutations (L452R-E484Q) in the B.1.617 variant on the binding of RBD to the host receptor ACE2. Our analysis revealed that the B.1.617 variant possesses different dynamic behaviours by altering dynamic-stability, residual flexibility and structural compactness. Moreover, the new variant had altered the bonding network and structural-dynamics properties significantly. MM/GBSA technique was used, which further established the binding differences between the wild type and B.1.617 variant. In conclusion, this study provides a strong impetus to develop novel drugs against the new SARS-CoV-2 variants.