Amino Acid Residues In Proteins Research Articles

Synthesis and anti-inflammatory activity of mogrol derivatives modified at C24 site

Mogrol, the aglycone of well-known sweeter mogrosides, shows potent anti-inflammatory activity. In this study, forty-two mogrol derivatives bearing various pharmacophores with oxygen or nitrogen atoms were designed and synthesized via structural modification at C24 site, and their anti-inflammatory activity were screened against lipopolysaccharide (LPS)-induced RAW264.7 cells. Compared with mogrol, most of derivatives exhibited stronger inhibition of NO production without cytotoxicity. In particular, compound B5 that contained an indole motif effectively suppressed the secretion of inflammatory mediators including TNF-α and IL-6, and inhibited the expression levels of TLR4, p-p65 and iNOS proteins. Molecular docking showed that the active B5 interacted with amino acid residues of iNOS protein through π-π stacking and hydrophobic interactions with binding affinity value of −12.1 kcal/mol, which was much stronger than mogrol (−8.9 kcal/mol). These results suggest that derivative B5 is a promising anti-inflammatory molecule and the strategy of hybridizing indole skeleton on mogrol is worthy for further attention.

Synthesis and evaluation of pentacyclic triterpenoids conjugates as novel HBV entry inhibitors targeting NTCP receptor

Chronic liver diseases caused by hepatitis B virus (HBV) are the accepted main cause leading to liver cirrhosis, hepatic fibrosis, and hepatic carcinoma. Sodium taurocholate cotransporting polypeptide (NTCP), a specific membrane receptor of hepatocytes for triggering HBV infection, is a promising target against HBV entry. In this study, pentacyclic triterpenoids (PTs) including glycyrrhetinic acid (GA), oleanolic acid (OA), ursolic acid (UA) and betulinic acid (BA) were modified via molecular hybridization with podophyllotoxin respectively, and resulted in thirty-two novel conjugates. The anti-HBV activities of conjugates were evaluated in HepG2.2.15 cells. The results showed that 66% of the conjugates exhibited lower toxicity to the host cells and had significant inhibitory effects on the two HBV antigens, especially HBsAg. Notably, the compounds BA-PPT1, BA-PPT3, BA-PPT4, and UA-PPT3 not only inhibited the secretion of HBsAg but also suppressed HBV DNA replication. A significant difference in the binding of active conjugates to NTCP compared to the HBV PreS1 antigen was observed by SPR assays. The mechanism of action was found to be the competitive binding of these compounds to the NTCP 157–165 epitopes, blocking HBV entry into host cells. Molecular docking results indicated that BA-PPT3 interacted with the amino acid residues of the target protein mainly through π-cation, hydrogen bond and hydrophobic interaction, suggesting its potential as a promising HBV entry inhibitor targeting the NTCP receptor.

Molecular characterization, immune functions and DNA protective effects of peroxiredoxin-1 gene in Antheraea pernyi

Peroxiredoxins are antioxidant proteins that detoxify peroxynitrite, hydrogen peroxide, and organic hydroperoxides, impacting various physiological processes such as immune responses, apoptosis, cellular homeostasis, and so on. In the present study, we identified and characterized peroxiredoxin 1 from Antheraea pernyi (thereafter designated as ApPrx-1) that encodes a predicted 195 amino acid residue protein with a 21.8 kDa molecular weight. Quantitative real-time PCR analysis revealed that the mRNA level of ApPrx-1 was highest in the hemocyte, fat body, and midgut. Immune-challenged larval fat bodies and hemocytes showed increased ApPrx-1 transcript. Moreover, ApPrx-1 expression was induced in hemocytes and the whole body of A. pernyi following exogenous H2O2 administration. A DNA cleavage assay performed using recombinant ApPrx-1 protein showed that rApPrx-1 protein manifests the ability to protect supercoiled DNA damage from oxidative stress. To test the rApPrx-1 protein antioxidant activity, the ability of the rApPrx-1 protein to remove H2O2 was assessed in vitro using rApPrx-1 protein and DTT, while BSA + DDT served as a control group. The results revealed that ApPrx-1 can efficiently remove H2O2 in vitro. In the loss of function analysis, we found that ApPrx-1 significantly increased the levels of H2O2 in ApPrx-1-depleted larvae compared to the control group. We also found a significantly lower survival rate in the larvae in which ApPrx-1 was knocked down. Interestingly, the antibacterial activity was significantly higher in the ApPrx-1 depleted larvae, compared to the control. Collectively, evidence strongly suggests that ApPrx-1 may regulate physiological activities and provides a reference for further studies to validate the utility of the key genes involved in reliving oxidative stress conditions and regulating the immune responses of insects.

Investigation of the Interaction of Monoamine Oxidase (MAO) Inhibitor Biscoumarins with Hemoglobin and Albumin Using Multi-Spectroscopic Techniques and Molecular Docking

Coumarin and their derivatives, such as biscoumarins, which are found as active ingredients in a variety of drugs, possess a wide range of biological activities. In this paper, the inhibitory effects of biscoumarin derivatives B1 and B2 on monoamine oxidase (MAO) enzyme activity were studied. Their binding mechanism to bovine serum hemoglobin (BHb) and bovine serum albumin (BSA) was investigated using molecular docking and multi-spectroscopic methods such as absorption, fluorescence, and synchronized fluorescence spectroscopy. The fluorescence quenching processes were analyzed using the Stern-Volmer, Lineweaver-Burk, and Hill methods. In this regard, Stern-Volmer quenching constants (Ksv), Lineweaver-Burk constants (KLB), and binding constants (Ka) were calculated at three temperatures (298, 310 and 322 K). The fluorescence studies showed that the binding of biscoumarins to BHb or BSA was a driven by static quenching. The Ka values of B2 to BHb and BSA (2179.11 and 18453.98 x 105 M-1) was larger than B1 at corresponding temperature (62.75 and 599.90 x105 M-1 for BHb and BSA, respectively), which indicates that the affinity of B2 toward the proteins were higher than that of B1. Moreover, thermodynamic parameters explained that hydrogen bonds were the main binding force stabilizing the protein-ligand complexes. Additionally, molecular docking studies have revealed binding energies, ligand efficiency values, and interactions with amino acid residues of proteins. The results of these investigations may be helpful in analyzing drug-protein binding of biscoumarins for potential future applications.

Effects of Glyprolins on Lipid and Protein Peroxidation in the Hypothalamic Region in Experimental Hyperthyroidism

We investigate effects of glyproline-type neuropeptides on lipid and protein peroxidation in the hypothalamic brain region of rats under the conditions of experimental hyperthyroidism. The state of hyperthyroidism in animals was simulated by intragastric administration of sodium pentahydrate of L-thyroxine at a dose of 150 µg/kg for 21 days. The following experimental groups (n=10) were formed: 1) control group — intact animals (control); 2) animals treated with L-thyroxine sodium salt pentahydrate (hyperthyroidism); 3) animals treated with Thr-Lys-Pro-Arg-Pro-Gly-Pro (celank); 4) animals treated with Pro-GlyPro (doses of 87 and 33 µg/kg/day, respectively) intraperitoneally daily during 21 days starting one day after the last administration of sodium pentahydrate of L-thyroxine. The level of lipid peroxidation processes was assessed by the initial level of TBA-reactive products, spontaneous and ascorbate-dependent lipid peroxidation rates in hypothalamic tissue homogenate. Protein peroxidation products were determined by the reaction between oxidized amino acid residues of proteins and 2,4-dinitrophenylhydrazone. The enzymatic part of the antioxidant system of hypothalamic region was estimated by measuring the activity of superoxide dismutase and catalase. In the setting of Thr-Lys-Pro-Arg-Pro-Gly-Pro and Pro-Gly-Pro administration under experimental hyperthyroidism, the intensity of lipid and protein peroxidation processes was decreased, while the activity of antioxidant enzymes-superoxide dismutase and catalase was restored in the hypothalamic tissue. The experimental data obtained indicate that, under the conditions of experimental hyperthyroidism, Thr-Lys-Pro-Arg-Pro-Gly-Pro and Pro-Gly-Pro compounds exhibit an antioxidant and antiradical activity with respect to parameters of lipoperoxidation and oxidative modification of proteins, as well as with respect to enzymatic defense systems in the hypothalamic brain region of laboratory animals.

Anti-Inflammatory Sesquiterpenes from Fruiting Bodies of Schizophyllum commune.

Schizophyllum commune, a fleshy fungus, is an important medicinal and food-homologous mushroom in China. In this work, eight undescribed sesquiterpenes schizomycins A-H (1-8) and one new meroterpenoid schizomycin I (9) together with three known analogues (10-12) were isolated from fruiting bodies of S. commune. Their planar structures were established by extensive spectroscopic and mass spectrometric data. The absolute configurations of compounds 1, 2, and 4 were determined by single crystal X-ray diffraction, and compounds 3 and 5-9 were confirmed by electronic circular dichroism calculations. Anti-inflammatory activities of all isolated compounds were evaluated for their inhibitory effects on IL-6 and IL-1β production in RAW 264.7 cells. Among them, compound 7 exhibited significant IL-6 inhibitory activity with an IC50 value of 3.6 μM. The results of molecular docking showed that compound 7 interacts with amino acid residues (Gly117, Lys118, Asp120, Thr166, and Try168) of the IL-6 receptor protein through hydrogen bonding.

EFFECT OF ACTG(4-7)-PRO-GLY-PRO AND ACTG(6-9)-PRO-GLY-PRO ON THE INTENSITY OF REDOX PROCESSES AND BEHAVIOURAL STATUS OF RATS UNDER THYROID HYPERFUNCTION CONDITIONS

The aim of the work is to study the effect of melanocortins on the intensity of redox processes in the prefrontal region of the brain and the behavioral status of rats against the background of thyroid hyperfunction. Material and methods. In the experiment, the test animals were divided into the following groups (n = 10): intact rats (control); animals treated with sodium salt pentahydrate L-thyroxine (hyperthyroidism); rats receiving ACTH (4-7)-Pro-Gly-Pro and individuals receiving ACTH (6-9)-Pro-Gly-Pro, re-spectively, intraperitoneally daily for 21 days, starting from day 22 of administration of sodium salt pentahydrate L-thyroxine. Hyperthyroidism was simulated in animals by intragastric administration of L-thyroxine sodium salt pentahydrate for 42 days. The intensity of lipoperoxidation processes was studied by the content of diene and triene conjugates in the prefrontal zone tissue, the initial level of TBА-reactive products, and the rates of spon-taneous and ascorbic peroxidation of lipids. The level of protein peroxidation products was determined by the reaction of oxidized amino acid residues of proteins with 2,4-dinitrophenyl hydrazone. Evaluation of the enzymatic unit of the antioxidant system of the prefrontal region was carried out by de-termining the activity of superoxide dismutase and catalase. The behavioral status of laboratory rodents was studied using the «Hole board» test. Results. The use of ACTH(4-7)-Pro-Gly-Pro and ACTH(6-9)-Pro-Gly-Pro compounds under experimental hyperthyroidism led to a decrease in the intensity of the processes of peroxidation of lipids, proteins and the restoration of levels of activity of antioxidant enzymes – superoxide dismutase and catalase in the tissue of the prefrontal region of the brain of animals, as well as correction behavioral indicators in the «Hole board» test. Conclusion. The obtained experimental data indicate that ACTH(4-7)-Pro-Gly-Pro and ACTH(6-9)-Pro-Gly-Pro show antiperoxy and antioxidant activity against lipoperoxidation parameters and oxidative modification of proteins and enzymatic protection systems in the prefrontal brain zone tissue, as well as exert psychocorregating action on behavioral responses of white rats against the background of thyroid hyperfunction.

2′-FL glycosylation of bovine milk protein matrices alters protein structure and reduces antibody affinity

Three glycosylated proteins were prepared from 2′-fucosyllactose (2′-FL) with α-lactalbumins, β-lactoglobulin and whey protein isolate, respectively, and their protein structures, glycosylation sites, digestion and transport capacities, and antibody binding capacities were determined. The findings indicate that following heating at 68 °C for 90 min, 2′-FL can form stable bonds with amino acid residues in milk proteins, leading to enhanced antioxidant properties, reduced hydrophobicity, and modified secondary structure content of the proteins. The glycosylation sites were mainly located in the amino acid sequence of β-lactoglobulin (K47, K91 and K135) and significantly reduce the affinity of its specific antibodies. 2′-FL glycosylation did not have a significant effect on the digestibility of cow's milk proteins, but it could significantly reduce the affinity of their specific antibody, indicating that it could be used in producing hypoallergenic dairy product.

Protein Target Prediction and Validation of Small Molecule Compound.

Proteins are fundamental to human physiology, with their targets being crucial in research and drug development. The identification and validation of crucial protein targets have become integral to drug development. Molecular docking is a computational tool widely utilized to investigate protein-ligand binding, especially in the context of drug and protein target interactions. For the experimental verification of the binding and to access the binding of the drug and its target directly, the cellular thermal shift assay (CETSA) method is used. This study aimed to integrate molecular docking with CETSA to predict and validate interactions between drugs and vital protein targets. Specifically, we predicted the interaction between xanthatin and Keap1 protein as well as its binding mode through molecular docking analysis, followed by verification of the interaction using the CETSA assay. Our results demonstrated that xanthatin could establish hydrogen bonds with specific amino acid residues of Keap1 protein and reduce the thermostability of Keap1 protein, indicating that xanthatin could directly interact with Keap1 protein.

Biochemical characterization and mutational analysis of the NurA protein from the hyperthermophilic euryarchaeon Thermococcus barophilus Ch5

Archaeal NurA protein plays a key role in producing 3′-single stranded DNA used for homologous recombination repair, together with HerA, Mre11, and Rad50. Herein, we describe biochemical characteristics and roles of key amino acid residues of the NurA protein from the hyperthermophilic euryarchaeon Thermococcus barophilus Ch5 (Tba-NurA). Tba-NurA possesses 5′–3′ exonuclease activity for degrading DNA, displaying maximum efficiency at 45oC ∼ 65oC and at pH 8.0 in the presence of Mn2+. The thermostable Tba-NurA also possesses endonuclease activity capable of nicking plasmid DNA and circular ssDNA. Mutational data demonstrate that residue D49 of Tba-NurA is essential for exonuclease activity and is involved in binding ssDNA since the D49A mutant lacked exonuclease activity and reduced ssDNA binding. The R96A and R129A mutants had no detectable dsDNA binding, suggesting that residues R96 and R129 are important for binding dsDNA. The abolished degradation activity and reduced dsDNA binding of the D120A mutant suggest that residue D120 is essential for degradation activity and dsDNA binding. Additionally, residues Y392 and H400 are important for exonuclease activity since these mutations resulted in exonuclease activity loss. To our knowledge, it is the first report on biochemical characterization and mutational analysis of the NurA protein from Thermococcus.

Protective effect of tretinoin on cervical cancer growth and proliferation through downregulation of pFAK2 expression.

Cervical cancer, a life-threatening disease, is the seventh most commonly detected cancer among women throughout the world. The present study investigated the effect of tretinoin on cervical cancer growth and metastasis in vitro and in vivo in the mice model. Cell Counting Kit-8, clonogenic survival, and transwell chamber assays were used for determination cells proliferation, colony formation, and invasiveness. Western blotting assay was used for assessment of protein expression whereas AutoDock Vina and Discovery studio software for in silico studies. Tretinoin treatment significantly (p < .05) reduced the proliferation of HT-3 and Caski cells in concentration-based manner. Incubation with tretinoin caused a significant decrease in clonogenic survival of HT-3 and Caski cells compared with the control cultures. The invasive potential of HT-3 cells was decreased to 18%, whereas that of Caski cells to 21% on treatment with 8 μM concentration of tretinoin. In HT-3 cells, tretinoin treatment led to a prominent reduction in p-focal adhesion kinase (FAK), matrix metalloproteinases (MMP)-2, and MMP-9 expression in HT-3 cells. Treatment of the cervical cancer mice model with tretinoin led to a prominent decrease in tumor growth. The metastasis of tumor in model cervical cancer mice group was effectively inhibited in spleen, intestines, and peritoneal cavity. In silico studies showed that tretinoin interacts with alanine, proline, isoleucine, and glycine amino acid residues of FAK protein to block its activation. The 2-dimensional diagram of interaction of tretinoin with FAK protein revealed that tretinoin binds to alanine and glycine amino acids through conventional hydrogen bonding. In summary, tretinoin suppressed the proliferation, colony formation, and invasiveness of cervical cancer cells in vitro. It decreased the expression of activated focal adhesion kinase, MMP-2, and MMP-9 in HT-3 cells in dose-dependent manner. In silico studies showed that tretinoin interacts with alanine and glycine amino acids through conventional hydrogen bonding. In vivo data demonstrated that treatment of the cervical cancer mice model with tretinoin led to a prominent decrease in tumor growth. Therefore, tretinoin can be developed as an effective therapeutic agent for cervical cancer treatment.

Molecular Mechanisms and the Significance of Synonymous Mutations.

Synonymous mutations result from the degeneracy of the genetic code. Most amino acids are encoded by two or more codons, and mutations that change a codon to another synonymous codon do not change the amino acid in the gene product. Historically, such mutations have been considered silent because they were assumed to have no to very little impact. However, research in the last few decades has produced several examples where synonymous mutations play important roles. These include optimizing expression by enhancing translation initiation and accelerating or decelerating translation elongation via codon usage and mRNA secondary structures, stabilizing mRNA molecules and preventing their breakdown before translation, and faulty protein folding or increased degradation due to enhanced ubiquitination and suboptimal secretion of proteins into the appropriate cell compartments. Some consequences of synonymous mutations, such as mRNA stability, can lead to different outcomes in prokaryotes and eukaryotes. Despite these examples, the significance of synonymous mutations in evolution and in causing disease in comparison to nonsynonymous mutations that do change amino acid residues in proteins remains controversial. Whether the molecular mechanisms described by which synonymous mutations affect organisms can be generalized remains poorly understood and warrants future research in this area.

Electrochemistry-enabled residue-specific modification of peptides and proteins.

Selective chemical reactions at precise amino acid residues of peptides and proteins have become an exploding field of research in the last few decades. With the emerging utility of bioconjugated peptides and proteins as drug leads and therapeutic agents, the design of smart protocols to modulate and conjugate biomolecules has become necessary. During this modification, the most important concern of biochemists is to keep intact the structural integrity of the biomolecules. Hence, a soft and selective biocompatible reaction environment is necessary. Electrochemistry, a mild and elegant tunable reaction platform to synthesize complex molecules while avoiding harsh and toxic chemicals, can provide such a reaction condition. However, this strategy is yet to be fully exploited in the field of selective modification of polypeptides. With this possibility, the use of electrochemistry as a reaction toolbox in peptide and protein chemistry is flourishing day by day. Unfortunately, there is no suitable review article summarizing the residue-specific modification of biomolecules. The present review provides a comprehensive summary of the latest manifested electrochemical approaches for the modulation of five redox-active amino acid residues, namely cysteine, tyrosine, tryptophan, histidine and methionine, found in peptides and proteins. The article also highlights the incredible potential of electrochemistry for the regio- as well as chemoselective bioconjugation strategy of biomolecules.

STUDY MOLECULES DOCKING OF ALKALOIDS IN KRATOM ON SEROTONIN TRANSPORTER (SERT), NOREPINEPHRINE TRANSPORTER (NET), AND MONOAMINE OXIDASE (MAO)

Kratom (Mitragyna speciosa Korth) is a tropical plant originating from Southeast Asia that predominantly contains alkaloid compounds and can potentially maintain levels of monoamine compounds in the body to treat depression. The study aimed to examine the potential of 8 alkaloid compounds in kratom as antidepressants towards four target proteins: Serotonin Transporter (SERT), Dopamine Transporter (DOPAT), Leucine Transporter (LEUT), and Monoamine Oxidase (MAO) via molecular docking. The Pyrx program is used with exhaustiveness 106 as the protocol, and the grid is adapted to the active site of each receptor. The affinity values ​​of the alkaloid compounds in kratom are mitragynine, 7-hydroxy mitragynine, speciociliatine, paynantheine, speciogynine, corynantheidine, mitraciliatine, and 9-hydroxycorynantheidine, for MAO were -7.1, -6.1, -5.7, -6.7, -5.7, -7.7, -5.7, and -5.7 kcal/ mole. All compounds bind to amino acid residues in the target protein through hydrogen and pi (π) bonds. All the tested alkaloid compounds have the potential to be re-uptake inhibitors SERT, DOPAT, LEUT, and Monoamine Oxidase (MAO).

Dual Effects of 3-epi-betulin from Daphniphyllum glaucescens in Suppressing SARS-CoV-2-Induced Inflammation and Inhibiting Virus Entry

The continuous emergence of SARS-CoV-2 variants has led to a protracted global COVID-19 pandemic with significant impacts on public health and global economy. While there are currently available SARS-CoV-2 vaccines and therapeutics, most of the FDA-approved antiviral agents directly target viral proteins. However, inflammation is the initial immune pathogenesis induced by SARS-CoV-2 infection, there is still a need to find additional agents that can control the virus in the early stages of infection to alleviate disease progression for the next pandemic. Here, we find that both the spike protein and its receptor CD147 are crucial for inducing inflammation by SARS-CoV-2 in THP-1 monocytic cells. Moreover, we find that 3-epi-betulin, isolated from Daphniphyllum glaucescens, reduces the level of proinflammatory cytokines induced by SARS-CoV-2, consequently resulting in a decreased viral RNA accumulation and plaque formation. In addition, 3-epi-betulin displays a broad-spectrum inhibition of entry of SARS-CoV-2 pseudoviruses, including Alpha (B.1.1.7), Eplison (B.1.429), Gamma (P1), Delta (B.1.617.2) and Omicron (BA.1). Moreover, 3-epi-betulin potently inhibits SARS-CoV-2 infection with an EC50 of &lt;20 μM in Calu-3 lung epithelial cells. Bioinformatic analysis reveals the chemical interaction between the 3-epi-betulin and the spike protein, along with the critical amino acid residues in the spike protein that contribute to the inhibitory activity of 3-epi-betulin against virus entry. Taken together, our results suggest that 3-epi-betulin exhibits dual effect: it reduces SARS-CoV-2-induced inflammation and inhibits virus entry, positioning it as a potential antiviral agent against SARS-CoV-2.

Metal-binding peptides and their potential to enhance the absorption and bioavailability of minerals.

Minerals including calcium, iron, zinc, magnesium, and copper have several human nutritional functions due to their metabolic activities. Body tissues require sufficient levels of a variety of micronutrients to maintain their health. To achieve these micronutrient needs, dietary consumption must be adequate. Dietary proteins may regulate the biological functions of the body in addition to acting as nutrients. Some peptides encoded in the native protein sequences are primarily responsible for the absorption and bioavailability of minerals in physiological functions. Metal-binding peptides (MBPs) were discovered as potential agents for mineral supplements. Nevertheless, sufficient studies on how MBPs affect the biological functions of minerals are lacking. The hypothesis is that the absorption and bioavailability of minerals are significantly influenced by peptides, and these properties are further enhanced by the configuration and attribute of the metal-peptide complex. In this review, the production of MBPs is discussed using various key parameters such as the protein sources and amino acid residues, enzymatic hydrolysis, purification, sequencing and synthesis and in silico analysis of MBPs. The mechanisms of metal-peptide complexes as functional food ingredients are elucidated, including metal-peptide ratio, precursors and ligands, complexation reaction, absorbability and bioavailability. Finally, the characteristics and application of different metal-peptide complexes are also described.

Mitophagy As a Novel Cell-Protective Mechanism Against Mitochondrial Ferroptosis in Acute Myeloid Leukemia

Background:Resistance to apoptosis poses a major clinical challenge in the therapy of acute myeloid leukemia (AML), leading to dismal clinical outcomes. Ferroptosis, another mode of regulated cell death that could bypass apoptosis resistance, has recently been investigaed for its therapeutic potential in therapy-resistant cancers. Meanwhile, accumulating evidence suggests that the regulation of ferroptosis is highly context-dependent and thus varies among cancer types. Hence, it is crucial to elucidate unique regulatory mechanisms in each tumor type for developing a therapeutic strategy. We recently found cell survival dependence of AML cells on GPX4, a major negative regulator of ferroptosis, and unique mechanistic involvement of mitochondria in AML ferroptosis. Results: We demonstrate that a selective GPX4 inhibitor ML210 as well as GPX4 knockdown induces ferroptosis, defined by iron- and lipid-peroxidation-dependence, in AML cell lines and primary AML samples, regardless of venetoclax resistance or genetic features including TP53 mutations. The cell sensitivity was negatively correlated with GPX4 protein expressions (i.e., the lower the expression, the higher the sensitivity). Normal bone marrow cells exhibited higher GPX4 levels compared to AML cells and showed significantly less sensitivity to ML210 treatment, suggesting a potential therapeutic window for GPX4 inhibition. Anti-leukemia effects of GPX4 knockdown were also observed in vivo, leading to survival prolongation in AML xenografted mice. Importantly, ferroptosis induction in AML cells is accompanied with and highly dependent on mitochondrial lipid peroxidation. 4-Hydroxynonenal (4-HNE) is one of the most biologically active products of lipid peroxidation, which is known to form adducts with certain amino acid residues of proteins to induce protein aggregates in the cytosol. We thus hypothesized that mitochondrial lipid peroxides induce mitochondrial protein aggregation in AML cell ferroptosis. Intriguingly, confocal microscopy revealed mitochondrial protein aggregation as evidenced by the co-localization of TOMM20 and protein aggregates detected by the PROTEOSTAT assay (Figure 1). Given that mitochondrial stress induced by protein aggregation reportedly induces mitophagy as a cellular stress response, we presumed that AML cells under the stress of ferroptosis may also show increased mitophagy. Indeed, GPX4 inhibition induced mitophagy, particularly in persistent AML cells, which was detected by the Mitophagy dye as mitochondrial acidification when fused with lysosomes, suggesting that mitophagy is a cell-protective response against ferroptosis as a quality control mechanism of malfunctioning mitochondria with lipid peroxidation and protein aggregation. Indeed, among persistent AML cells exposed to GPX4 inhibition, cells with more mitophagy were more resistant to further treatment with ML210 compared to those with less mitophagy. Consistently, an analogue of the ceramide-mediated mitophagy inducer LCL-461 (Dany, et al. 2016 Blood) suppressed GPX4 inhibition-induced ferroptosis in AML cells. On the other hand, inhibition of ULK1-dependent mitophagy through ULK1 knockout or the ULK1 inhibitor SBI0206965 (Bhattacharya, et al. 2021 Molecular Cancer Research) synergistically enhanced the anti-leukemia effects of GPX4 inhibition (Figure 2), suggesting a more effective therapeutic potential in ferroptosis in AML. Conclusion:Our study revealed that ferroptosis in AML cells causes mitochondrial protein aggregation and that mitophagy is induced in ferroptosis-persistent AML cells as a cell-protective mechanism against ferroptosis. This provides novel mechanistic insights into mitochondrial ferroptosis in AML, suggesting the therapeutic potential of co-targeting mitophagy in ferroptosis-based therapy. Further studies are in progress to elucidate the molecular mechanisms involved as well as the in vivo efficacy and safety of the combinatorial treatment.

Predicting pKa of the carboxylic acid group in water solutions of amino acids based on molecular structures using machine learning QSPR methods

Amino acids as biomolecules are essential building blocks of proteins, which are fundamental to life and living organisms. The pKa values of amino acid residues in proteins can impact their structure and function. Furthermore, the pKa can be used to optimize experimental conditions for protein purification. In this research, a data set containing the pKa of the carboxylic acid group for 52 amino acids from various families in a water solution was used. The quantity structure property relationship (QSPR) approach was applied in our work for predicting the pKa using different machine learning methods. Initially, the amine molecules were drawn, and optimized and 3224 molecular descriptors were calculated for each molecule. Selecting among molecular descriptors using genetic algorithm-multi linear regression (GA-MLR) shows that the model with 8 descriptors is the best economic one. The comparison of results shows that the feed forward neural network (FFNN) is better than the particle swarm optimization-support vector machine (PSO-SVM) model. The coefficients of determination at different models follow the order of R2 FFNN= 0.9987 > R2 PSO-SVM= 0.9971 > R2 GA-MLR= 0.9951. The R2test for FFNN and PSO-SVM are near the R2train and this evidence proves the validation of models. Consequently, by using FFNN, the pKa of the carboxylic acid group in water solutions of amino acids can be predicted without any experimental costs with a low average absolute relative error (AARE%) of 2.82.

Biophysical reviews top five: voltage-dependent charge movement in nerve and muscle.

The discovery of gating currents and asymmetric charge movement in the early 1970s represented a remarkable leap forward in our understanding of the biophysical basis of voltage-dependent events that underlie electrical signalling that is vital for nerve and muscle function. Gating currents and charge movement reflect a fundamental process in which charged amino acid residues in an ion channel protein move in response to a change in the membrane electrical field and therefore activate the specific voltage-dependent response of that protein. The detection of gating currents and asymmetric charge movement over the past 50 years has been pivotal in unraveling the multiple molecular and intra-molecular processes which lead to action potentials in excitable tissues and excitation-contraction (EC) coupling in skeletal muscle. The recording of gating currents and asymmetric charge movement remains an essential component of investigations into the basic molecular mechanisms of neuronal conduction and muscle contraction.

Unique Patterns in Amino Acid Sequences of Aging-Related Proteins.

Aging has strong genetic components and the list of genes that may regulate the aging process is collected in the GenAge database. There may be characteristic patterns in the amino acid sequences of aging-related proteins that distinguish them from other proteins and this information will lead to a better understanding of the aging process. To test this hypothesis, human protein sequences are extracted from the UniProt database and the relative frequency of every amino acid residue in aging-related proteins and the remaining proteins is calculated. The main observation is that the mean relative frequency of aspartic acid (D) is consistently higher, while the mean relative frequencies of tryptophan (W) and leucine (L) are consistently lower in aging-related proteins compared to the non-aging-related proteins for the human and four examined model organisms. It is also observed that the mean relative frequency of aspartic acid is higher, while the mean relative frequency of tryptophan is lower in pro-longevity proteins compared to anti-longevity proteins in model organisms. Finally, it is found that aging-related proteins tend to be longer than non-aging-related proteins. It is hoped that this analysis initiates further computational and experimental research to explore the underlying mechanisms of these findings.