Showing papers in "Biochemistry in 2017"
TL;DR: This guide will help triage candidate methods for peptide alteration and will serve as a starting point for those seeking to solve long-standing challenges.
Abstract: Advances in bioconjugation and native protein modification are appearing at a blistering pace, making it increasingly time consuming for practitioners to identify the best chemical method for modifying a specific amino acid residue in a complex setting. The purpose of this perspective is to provide an informative, graphically rich manual highlighting significant advances in the field over the past decade. This guide will help triage candidate methods for peptide alteration and will serve as a starting point for those seeking to solve long-standing challenges.
320 citations
TL;DR: The enzymology community needs to embrace the essential role of web tools that allow the protein and genome sequence databases to be leveraged for novel insights into enzymological problems.
Abstract: The exponentially increasing number of protein and nucleic acid sequences provides opportunities to discover novel enzymes, metabolic pathways, and metabolites/natural products, thereby adding to our knowledge of biochemistry and biology. The challenge has evolved from generating sequence information to mining the databases to integrating and leveraging the available information, i.e., the availability of “genomic enzymology” web tools. Web tools that allow identification of biosynthetic gene clusters are widely used by the natural products/synthetic biology community, thereby facilitating the discovery of novel natural products and the enzymes responsible for their biosynthesis. However, many novel enzymes with interesting mechanisms participate in uncharacterized small-molecule metabolic pathways; their discovery and functional characterization also can be accomplished by leveraging information in protein and nucleic acid databases. This Perspective focuses on two genomic enzymology web tools that assis...
161 citations
TL;DR: The contribution of individual proteins and macromolecular complexes in cell mechanics and the tools that make it possible to quantitatively decipher the biochemical machinery that contributes to bacterial cell mechanics are discussed.
Abstract: Cellular mechanical properties play an integral role in bacterial survival and adaptation. Historically, the bacterial cell wall and, in particular, the layer of polymeric material called the peptidoglycan were the elements to which cell mechanics could be primarily attributed. Disrupting the biochemical machinery that assembles the peptidoglycan (e.g., using the β-lactam family of antibiotics) alters the structure of this material, leads to mechanical defects, and results in cell lysis. Decades after the discovery of peptidoglycan-synthesizing enzymes, the mechanisms that underlie their positioning and regulation are still not entirely understood. In addition, recent evidence suggests a diverse group of other biochemical elements influence bacterial cell mechanics, may be regulated by new cellular mechanisms, and may be triggered in different environmental contexts to enable cell adaptation and survival. This review summarizes the contributions that different biomolecular components of the cell wall (e.g...
154 citations
TL;DR: The structural basis for family A GPCR activation is reviewed, with a focus on GPCRs for which structures are available in both active or active-like states and inactive states, and new technologies that may allow unanswered questions to be resolved are highlighted.
Abstract: G protein-coupled receptors (GPCRs) are critical regulators of human physiology and make up the largest single class of therapeutic drug targets. Although GPCRs regulate highly diverse physiology, they share a common signaling mechanism whereby extracellular stimuli induce conformational changes in the receptor that enable activation of heterotrimeric G proteins and other intracellular effectors. Advances in GPCR structural biology have made it possible to examine ligand-induced GPCR activation at an unprecedented level of detail. Here, we review the structural basis for family A GPCR activation, with a focus on GPCRs for which structures are available in both active or active-like states and inactive states. Crystallographic and other biophysical data show how chemically diverse ligands stabilize highly conserved conformational changes on the intracellular side of the receptors, allowing many different extracellular stimuli to utilize shared downstream signaling molecules. Finally, we discuss the remaini...
125 citations
TL;DR: This study informs a mechanism for electron bifurcation, offering insight into a unique method for delivery of low-potential electrons required for energy-intensive biochemical conversions.
Abstract: The biological reduction of dinitrogen (N2) to ammonia (NH3) by nitrogenase is an energetically demanding reaction that requires low-potential electrons and ATP; however, pathways used to deliver the electrons from central metabolism to the reductants of nitrogenase, ferredoxin or flavodoxin, remain unknown for many diazotrophic microbes. The FixABCX protein complex has been proposed to reduce flavodoxin or ferredoxin using NADH as the electron donor in a process known as electron bifurcation. Herein, the FixABCX complex from Azotobacter vinelandii was purified and demonstrated to catalyze an electron bifurcation reaction: oxidation of NADH (Em = −320 mV) coupled to reduction of flavodoxin semiquinone (Em = −460 mV) and reduction of coenzyme Q (Em = 10 mV). Knocking out fix genes rendered Δrnf A. vinelandii cells unable to fix dinitrogen, confirming that the FixABCX system provides another route for delivery of electrons to nitrogenase. Characterization of the purified FixABCX complex revealed the presenc...
116 citations
TL;DR: The observed increase in linker stiffness with reduced glycine content is much less pronounced than that predicted by a classical model developed by Flory and co-workers, and possible reasons for this discrepancy are discussed.
Abstract: Flexible polypeptide linkers composed of glycine and serine are important components of engineered multidomain proteins. We have previously shown that the conformational properties of Gly-Gly-Ser repeat linkers can be quantitatively understood by comparing experimentally determined Forster resonance energy transfer (FRET) efficiencies of ECFP-linker-EYFP proteins to theoretical FRET efficiencies calculated using wormlike chain and Gaussian chain models. Here we extend this analysis to include linkers with different glycine contents. We determined the FRET efficiencies of ECFP-linker-EYFP proteins with linkers ranging in length from 25 to 73 amino acids and with glycine contents of 33.3% (GSSGSS), 16.7% (GSSSSSS), and 0% (SSSSSSS). The FRET efficiency decreased with an increasing linker length and was overall lower for linkers with less glycine. Modeling the linkers using the WLC model revealed that the experimentally observed FRET efficiencies were consistent with persistence lengths of 4.5, 4.8, and 6.2 A for the GSSGSS, GSSSSS, and SSSSSS linkers, respectively. The observed increase in linker stiffness with reduced glycine content is much less pronounced than that predicted by a classical model developed by Flory and co-workers. We discuss possible reasons for this discrepancy as well as implications for using the stiffer linkers to control the effective concentrations of connected domains in engineered multidomain proteins.
115 citations
TL;DR: There is a strong inverse correlation between the time to form amyloid and the extent of vesicle leakage, and the possibility that the loss of phosphatidylserine asymmetry enhances IAPP amyloids formation and membrane damage in vivo via a positive feedback loop.
Abstract: Amyloid formation by islet amyloid polypeptide (IAPP) contributes to β-cell dysfunction in type 2 diabetes. Perturbation of the β-cell membrane may contribute to IAPP-induced toxicity. We examine the effects of lipid composition, salt, and buffer on IAPP amyloid formation and on the ability of IAPP to induce leakage of model membranes. Even low levels of anionic lipids promote amyloid formation and membrane permeabilization. Increasing the percentage of the anionic lipids, 1-palmitoyl-2-oleoyl-sn-glycero-3-phospho-l-serine (POPS) or 1,2-dioleoyl-sn-glycero-3-phospho(1′-rac-glycerol), enhances the rate of amyloid formation and increases the level of membrane permeabilization. The choice of zwitterionic lipid has no noticeable effect on membrane-catalyzed amyloid formation but in most cases affects leakage, which tends to decrease in the following order: 1,2-dioleoyl-sn-glycero-3-phosphocholine > 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine > sphingomyelin. Uncharged lipids that increase the level of me...
109 citations
TL;DR: This review will highlight the recent progress in the determination of the three-dimensional structures of these enzymes, their chemical mechanisms, and insights into their suitability as targets for the development of antibacterial agents.
Abstract: The eight enzymes responsible for the biosynthesis of the three branched-chain amino acids (l-isoleucine, l-leucine, and l-valine) were identified decades ago using classical genetic approaches based on amino acid auxotrophy. This review will highlight the recent progress in the determination of the three-dimensional structures of these enzymes, their chemical mechanisms, and insights into their suitability as targets for the development of antibacterial agents. Given the enormous rise in bacterial drug resistance to every major class of antibacterial compound, there is a clear and present need for the identification of new antibacterial compounds with nonoverlapping targets to currently used antibacterials that target cell wall, protein, mRNA, and DNA synthesis.
108 citations
TL;DR: The current understanding of metabolic enzyme complexes is discussed by primarily focusing on central carbon metabolism and closely associated metabolic pathways in a variety of organisms, as well as their regulation and functional contributions to cells.
Abstract: The organization of metabolic multienzyme complexes has been hypothesized to benefit metabolic processes and provide a coordinated way for the cell to regulate metabolism. Historically, their existence has been supported by various in vitro techniques. However, it is only recently that the existence of metabolic complexes inside living cells has come to light to corroborate this long-standing hypothesis. Indeed, subcellular compartmentalization of metabolic enzymes appears to be widespread and highly regulated. On the other hand, it is still challenging to demonstrate the functional significance of these enzyme complexes in the context of the cellular milieu. In this review, we discuss the current understanding of metabolic enzyme complexes by primarily focusing on central carbon metabolism and closely associated metabolic pathways in a variety of organisms, as well as their regulation and functional contributions to cells.
106 citations
TL;DR: Results suggest that GAGs may be utilized as attachment factors for host cell entry of Zika virus as they do in other pathogenic flaviviruses.
Abstract: In February 2016, the World Health Organization declared a Public Health Emergency of International Concern on Zika Virus (ZIKV), because of its association with severe fetal anomalies of congenitally infected humans. This has led to urgent efforts by academic, federal, and industry research groups to improve our understanding of the pathogenesis of ZIKV and to develop detection methods, therapeutic strategies, and vaccines. Although we still do not have the entire picture of the pathogenesis of ZIKV, extensive research has been conducted on related pathogenic flaviviruses (i.e., dengue virus, West Nile virus, and yellow fever virus). Binding to glycosaminoglycans (GAGs) through its envelope protein is the first step in successful host cell invasion of dengue virus. In this study, we examined ZIKV envelope protein (ZIKV E) binding to GAGs in a real time interaction study using surface plasmon resonance (SPR) to explore the role of GAGs in host cell entry of ZIKV into placenta and brain. ZIKV E strongly bi...
100 citations
TL;DR: In this paper, the authors review aggregation mechanisms of proteins with expanded polyglutamine (polyQ)tracts, structural consequences of expanded polyQ ranging from monomers to fibrillar aggregates, the impact of protein context and posttranslational modifications on aggregation, and a potential role for lipid membranes in aggregation.
Abstract: Several hereditary neurological and neuromuscular diseases are caused by an abnormal expansion of trinucleotide repeats. To date, there have been 10 of these trinucleotide repeat disorders associated with an expansion of the codon CAG encoding glutamine (Q). For these polyglutamine (polyQ) diseases, there is a critical threshold length of the CAG repeat required for disease, and further expansion beyond this threshold is correlated with age of onset and symptom severity. PolyQ expansion in the translated proteins promotes their self-assembly into a variety of oligomeric and fibrillar aggregate species that accumulate into the hallmark proteinaceous inclusion bodies associated with each disease. Here, we review aggregation mechanisms of proteins with expanded polyQ-tracts, structural consequences of expanded polyQ ranging from monomers to fibrillar aggregates, the impact of protein context and post-translational modifications on aggregation, and a potential role for lipid membranes in aggregation. As the p...
TL;DR: In the present review, the results of the own research as well as data provided in the corresponding literature on the interaction of brain-derived neurotrophic factor (BDNF) and glial cell line- derived neurotrophic factors (GDNF) with the 5-HT-system of the brain are considered.
Abstract: Neurotrophic factors play a key role in development, differentiation, synaptogenesis, and survival of neurons in the brain as well as in the process of their adaptation to external influences. The serotonergic (5-HT) system is another major factor in the development and neuroplasticity of the brain. In the present review, the results of our own research as well as data provided in the corresponding literature on the interaction of brain-derived neurotrophic factor (BDNF) and glial cell line-derived neurotrophic factor (GDNF) with the 5-HT-system of the brain are considered. Attention is given to comparison of BDNF and GDNF, the latter belonging to a different family of neurotrophic factors and being mainly considered as a dopaminergic system controller. Data cited in this review show that: (i) BDNF and GDNF interact with the 5-HT-system of the brain through feedback mechanisms engaged in autoregulation of the complex involving 5-HT-system and neurotrophic factors; (ii) GDNF, as well as BDNF, stimulates the growth of 5-HT neurons and affects the expression of key genes of the brain 5-HT-system–those coding tryptophan hydroxylase-2 and 5-HT1A and 5-HT2A receptors. In turn, 5-HT affects the expression of genes that control BDNF and GDNF in brain structures; (iii) the difference between BDNF and GDNF is manifested in different levels and relative distribution of expression of these factors in brain structures (BDNF expression is highest in hippocampus and cortex, GDNF expression in the striatum), in varying reaction of 5-HT2A receptors on BDNF and GDNF administration, and in different effects on certain types of behavior.
TL;DR: The recent advances in biochemistry, protein engineering, and organic synthesis that have allowed a triumphant return of chemical fluorophores to modern biological imaging are reviewed.
Abstract: Chemistry, once king of fluorescence microscopy, was usurped by the field of fluorescent proteins. The increased demands of modern microscopy techniques on the “photon budget” require better and brighter fluorophores, causing a renewed interest in synthetic dyes. Here, we review the recent advances in biochemistry, protein engineering, and organic synthesis that have allowed a triumphant return of chemical fluorophores to modern biological imaging.
TL;DR: Surprisingly, each of the two stem-loop structures that comprise the mini-ykkC motif appears to directly bind free guanidine in a cooperative manner, revealing that bacteria make extensive use of diverse Guanidine-responsive riboswitches to overcome the toxic effects of this compound.
Abstract: Recently, it was determined that representatives of the riboswitch candidates called ykkC and mini-ykkC directly bind free guanidine. These riboswitches regulate the expression of genes whose protein products are implicated in overcoming the toxic effects of this ligand. Thus, the relevant ykkC motif and mini-ykkC motif RNAs have been classified as guanidine-I and guanidine-II riboswitch RNAs, respectively. Moreover, we had previously noted that a third candidate riboswitch class, called ykkC-III, was associated with a distribution of genes similar to those of the other two motifs. Therefore, it was predicted that ykkC-III motif RNAs would sense and respond to the same ligand. In this report, we present biochemical data supporting the hypothesis that ykkC-III RNAs represent a third class of guanidine-sensing RNAs called guanidine-III riboswitches. Members of the guanidine-III riboswitch class bind their ligand with an affinity similar to that observed for members of the other two classes. Notably, there a...
TL;DR: Understanding the functional role of cytokines in cellular mechanisms of information processing and storage in the brain would allow developing preventive and therapeutic means for the treatment of neuropathologies related to impairment of these mechanisms.
Abstract: After accumulation of data showing that resident brain cells (neurons, astrocytes, and microglia) produce mediators of the immune system, such as cytokines and their receptors under normal physiological conditions, a critical need emerged for investigating the role of these mediators in cognitive processes. The major problem for understanding the functional role of cytokines in the mechanisms of synaptic plasticity, de novo neurogenesis, and learning and memory is the small number of investigated cytokines. Existing concepts are based on data from just three proinflammatory cytokines: interleukin-1 beta, interleukin-6, and tumor necrosis factor-alpha. The amount of information in the literature on the functional role of antiinflammatory cytokines in the mechanisms of synaptic plasticity and cognitive functions of mature mammalian brain is dismally low. However, they are of principle importance for understanding the mechanisms of local information processing in the brain, since they modulate the activity of individual cells and local neural networks, being able to reconstruct the processes of synaptic plasticity and intercellular communication, in general, depending on the local ratio of the levels of different cytokines in certain areas of the brain. Understanding the functional role of cytokines in cellular mechanisms of information processing and storage in the brain would allow developing preventive and therapeutic means for the treatment of neuropathologies related to impairment of these mechanisms.
TL;DR: Recent advances in small-molecule fluorescent indicators that rely on photoinduced electron transfer to sense voltage as well as refinements of voltage-sensitive fluorescent proteins and new opsin-based strategies for monitoring voltage changes are surveyed.
Abstract: Optical methods for interrogating membrane potential changes in neurons promise to revolutionize our ability to dissect the activity of individual cells embedded in neural circuits underlying behavior and sensation. A number of voltage imaging strategies have emerged in the past few years. This Perspective discusses developments in both small-molecule and genetically encoded fluorescent indicators of membrane potential. We survey recent advances in small-molecule fluorescent indicators that rely on photoinduced electron transfer to sense voltage as well as refinements of voltage-sensitive fluorescent proteins and new opsin-based strategies for monitoring voltage changes. We compare the requirements of fluorescent voltage indicators to those for more canonical Ca2+ sensing as a way to illuminate the particular challenges associated with voltage imaging.
TL;DR: Biophysical studies of the interaction of these two ferredoxins with other proteins involved in mitochondrial iron–sulfur cluster assembly show that both FDX1 and FDX2 can function in Fe–S cluster assembly in vitro.
Abstract: Ferredoxins play an important role as an electron donor in iron–sulfur (Fe–S) cluster biosynthesis. Two ferredoxins, human mitochondrial ferredoxin 1 (FDX1) and human mitochondrial ferredoxin 2 (FDX2), are present in the matrix of human mitochondria. Conflicting results have been reported regarding their respective function in mitochondrial iron–sulfur cluster biogenesis. We report here biophysical studies of the interaction of these two ferredoxins with other proteins involved in mitochondrial iron–sulfur cluster assembly. Results from nuclear magnetic resonance spectroscopy show that both FDX1 and FDX2 (in both their reduced and oxidized states) interact with the protein complex responsible for cluster assembly, which contains cysteine desulfurase (NFS1), ISD11 (also known as LYRM4), and acyl carrier protein (Acp). In all cases, ferredoxin residues close to the Fe–S cluster are involved in the interaction with this complex. Isothermal titration calorimetry results showed that FDX2 binds more tightly to ...
TL;DR: Current approaches in HCC targeted therapy are aimed at creating efficient strategies for interrupting tumor interactions with the stroma, as well as new developments in antitumor drug design are discussed.
Abstract: This review summarizes recently published data on the mechanisms of tumor cell interaction with the tumor microenvironment. Tumor stroma influences the processes of hepatocarcinogenesis, epithelial-to-mesenchymal transition, invasion, and metastasis. The tumor microenvironment includes both cellular and noncellular components. Main cellular components of hepatocellular carcinoma (HCC) stroma are tumor-associated fibroblasts, hepatic stellate cells, immune cells, and endothelial cells that produce extracellular components of tumor microenvironment such as extracellular matrix, various proteins, proteolytic enzymes, growth factors, and cytokines. The noncellular components of the stroma modulate signaling pathways in tumor cells and stimulate invasion and metastasis. The tumor microenvironment composition and organization can serve as prognostic factors in HCC pathogenesis. Current approaches in HCC targeted therapy are aimed at creating efficient strategies for interrupting tumor interactions with the stroma. Recent data on the composition and role of the microenvironment in HCC pathogenesis, as well as new developments in antitumor drug design are discussed.
TL;DR: This review focuses on the function of each member, interrelation among members, and the influence of manipulators and repressors in the ICE–CBF–COR pathway, which is thought to be rather important for cold resistance of plants.
Abstract: Low temperature is an abiotic stress that adversely affects the growth and production of plants. Resistance and adaptation of plants to cold stress is dependent upon the activation of molecular networks and pathways involved in signal transduction and the regulation of cold-stress related genes. Because it has numerous and complex genes, regulation factors, and pathways, research on the ICE-CBF-COR signaling pathway is the most studied and detailed, which is thought to be rather important for cold resistance of plants. In this review, we focus on the function of each member, interrelation among members, and the influence of manipulators and repressors in the ICE-CBF-COR pathway. In addition, regulation and signal transduction concerning plant hormones, circadian clock, and light are discussed. The studies presented provide a detailed picture of the ICE-CBF-COR pathway.
TL;DR: The articles in this issue are focused on different “hot points” in the research area of biochemical mechanisms supporting neuroplasticity.
Abstract: Biochemical processes in synapses and other neuronal compartments underlie neuroplasticity (functional and structural alterations in the brain enabling adaptation to the environment, learning, memory, as well as rehabilitation after brain injury). This basic molecular level of brain plasticity covers numerous specific proteins (enzymes, receptors, structural proteins, etc.) participating in many coordinated and interacting signal and metabolic processes, their modulation forming a molecular basis for brain plasticity. The articles in this issue are focused on different "hot points" in the research area of biochemical mechanisms supporting neuroplasticity.
TL;DR: The recent advances and strategies for designing such synthetic riboswitches based on natural or artificial components and readout systems are discussed, from trial-and-error approaches to rational design strategies.
Abstract: In synthetic biology, metabolic engineering, and gene therapy, there is a strong demand for orthogonal or externally controlled regulation of gene expression. Here, RNA-based regulatory devices represent a promising emerging alternative to proteins, allowing a fast and direct control of gene expression, as no synthesis of regulatory proteins is required. Besides programmable ribozyme elements controlling mRNA stability, regulatory RNA structures in untranslated regions are highly interesting for engineering approaches. Riboswitches are especially well suited, as they show a modular composition of sensor and response elements, allowing a free combination of different modules in a plug-and-play-like mode. The sensor or aptamer domain specifically interacts with a trigger molecule as a ligand, modulating the activity of the adjacent response domain that controls the expression of the genes located downstream, in most cases at the level of transcription or translation. In this review, we discuss the recent ad...
TL;DR: The generation and characterization of a set of recombinant antibody-like ADP-ribose binding proteins, in which naturally occurring ADPR binding domains, including macrodomains and WWE domains, have been functionalized by fusion to the Fc region of rabbit immunoglobulin.
Abstract: ADP-ribosylation is an enzyme-catalyzed post-translational modification of proteins in which the ADP-ribose (ADPR) moiety of NAD+ is transferred to a specific amino acid in a substrate protein. The biological functions of ADP-ribosylation are numerous and diverse, ranging from normal physiology to pathological conditions. Biochemical and cellular studies of the diverse forms and functions of ADPR require immunological reagents that can be used for detection and enrichment. The lack of a complete set of tools that recognize all forms of ADPR [i.e., mono-, oligo-, and poly(ADP-ribose)] has hampered progress. Herein, we describe the generation and characterization of a set of recombinant antibody-like ADP-ribose binding proteins, in which naturally occurring ADPR binding domains, including macrodomains and WWE domains, have been functionalized by fusion to the Fc region of rabbit immunoglobulin. These reagents, which collectively recognize all forms of ADPR with different specificities, are useful in a broad...
TL;DR: Two further series of peptides are constructed with a sequence similar to those of the KIA peptides, but with a constant charge of +7 for all lengths from 14 to 28 amino acids, indicating that charge has only a minor influence on activity.
Abstract: Hydrophobic mismatch is important for pore-forming amphipathic antimicrobial peptides, as demonstrated recently [Grau-Campistany, A., et al. (2015) Sci. Rep. 5, 9388]. A series of different length peptides have been generated with the heptameric repeat sequence KIAGKIA, called KIA peptides, and it was found that only those helices sufficiently long to span the hydrophobic thickness of the membrane could induce leakage in lipid vesicles; there was also a clear length dependence of the antimicrobial and hemolytic activities. For the original KIA sequences, the cationic charge increased with peptide length. The goal of this work is to examine whether the charge also has an effect on activity; hence, we constructed two further series of peptides with a sequence similar to those of the KIA peptides, but with a constant charge of +7 for all lengths from 14 to 28 amino acids. For both of these new series, a clear length dependence similar to that of KIA peptides was observed, indicating that charge has only a mi...
TL;DR: A new discovery in the C-terminal prion-like region of TDP-43 provides further support for the theory that liquid droplets such as stress granules may be precursors to pathological aggregates, linking environmental effectssuch as stress to the potential etiology of the disease.
Abstract: TDP-43 is a dimeric nuclear protein that plays a central role in RNA metabolism. In recent years, this protein has become a focal point of research in the amyotrophic lateral sclerosis and frontotemporal dementia (ALS/FTD) disease spectrum, as pathognomonic inclusions within affected neurons contain post-translationally modified TDP-43. A key question in TDP-43 research involves determining the mechanisms and triggers that cause TDP-43 to form pathological aggregates. This review gives a brief overview of the physiological and pathological roles of TDP-43 and focuses on the structural features of its protein domains and how they may contribute to normal protein function and to disease. A special emphasis is placed on the C-terminal prion-like region thought to be implicated in pathology, as it is where nearly all ALS/FTD-associated mutations reside. Recent structural studies of this domain revealed its crucial role in the formation of phase-separated liquid droplets through a partially populated α-helix. ...
TL;DR: The functions of GAPDH associated with DNA repair are discussed in detail and the mechanism of its translocation to the nucleus is not fully understood.
Abstract: Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is widely known as a glycolytic enzyme. Nevertheless, various functions of GAPDH have been found that are unrelated to glycolysis. Some of these functions presume interaction of GAPDH with DNA, but the mechanism of its translocation to the nucleus is not fully understood. When in the nucleus, GAPDH participates in the initiation of apoptosis and transcription of genes involved in antiapoptotic pathways and cell proliferation and plays a role in the regulation of telomere length. Several authors have shown that GAPDH displays the uracil-DNA glycosylase activity and interacts with some types of DNA damages, such as apurinic/apyrimidinic sites, nucleotide analogs, and covalent DNA adducts with alkylating agents. Moreover, GAPDH can interact with proteins participating in DNA repair, such as APE1, PARP1, HMGB1, and HMGB2. In this review, the functions of GAPDH associated with DNA repair are discussed in detail.
TL;DR: The classification of enteroviruses, the most common enterovirus infections and their clinical man- ifestations are mentioned, the current state of development of anti-enteroviral drugs are considered, and the viral Internal Ribosome Entry Site (IRES) is examined.
Abstract: The genus Enterovirus combines a portion of small (+)ssRNA-containing viruses and is divided into 10 species of true enteroviruses and three species of rhinoviruses. These viruses are causative agents of the widest spectrum of severe and deadly epidemic diseases of higher vertebrates, including humans. Their ubiquitous distribution and high pathogenici- ty motivate active search to counteract enterovirus infections. There are no sufficiently effective drugs targeted against enteroviral diseases, thus treatment is reduced to supportive and symptomatic measures. This makes it extremely urgent to develop drugs that directly affect enteroviruses and hinder their development and spread in infected organisms. In this review, we cover the classification of enteroviruses, mention the most common enterovirus infections and their clinical man- ifestations, and consider the current state of development of anti-enteroviral drugs. One of the most promising targets for such antiviral drugs is the viral Internal Ribosome Entry Site (IRES). The classification of these elements of the viral mRNA translation system is also examined.
TL;DR: The ability of a noncatalytic cysteine to react with a diverse set of electrophiles is demonstrated and the importance of proper spatial arrangements between a covalent inhibitor and its intended nucleophile is emphasized.
Abstract: There is growing interest in reversible and irreversible covalent inhibitors that target noncatalytic amino acids in target proteins. With a goal of targeting oncogenic K-Ras variants (e.g., G12D) by expanding the types of amino acids that can be targeted by covalent inhibitors, we survey a set of electrophiles for their ability to label carboxylates. We functionalized an optimized ligand for the K-Ras switch II pocket with a set of electrophiles previously reported to react with carboxylates and characterized the ability of these compounds to react with model nucleophiles and oncogenic K-Ras proteins. Here, we report that aziridines and stabilized diazo groups preferentially react with free carboxylates over thiols. Although we did not identify a warhead that potently labels K-Ras G12D, we were able to study the interactions of many electrophiles with K-Ras, as most of the electrophiles rapidly label K-Ras G12C. We characterized the resulting complexes by crystallography, hydrogen/deuterium exchange, and...
TL;DR: Advances in understanding vector immunity can be leveraged to develop novel control strategies for reducing the rate of transmission of both ancient and emerging threats to global health.
Abstract: Arthropods, especially ticks and mosquitoes, are the vectors for a number of parasitic and viral human diseases, including malaria, sleeping sickness, Dengue, and Zika, yet arthropods show tremendous individual variation in their capacity to transmit disease. A key factor in this capacity is the group of genetically encoded immune factors that counteract infection by the pathogen. Arthropod-specific pattern recognition receptors and protease cascades detect and respond to infection. Proteins such as antimicrobial peptides, thioester-containing proteins, and transglutaminases effect responses such as lysis, phagocytosis, melanization, and agglutination. Effector responses are initiated by damage signals such as reactive oxygen species signaling from epithelial cells and recognized by cell surface receptors on hemocytes. Antiviral immunity is primarily mediated by siRNA pathways but coupled with interferon-like signaling, antimicrobial peptides, and thioester-containing proteins. Molecular mechanisms of imm...
TL;DR: A combined solution nuclear magnetic resonance spectroscopic and computational investigation demonstrates that an interstrand halogen bond is capable of conformational stabilization of a β-hairpin foldamer comparable to an analogous hydrogen bond.
Abstract: Halogen bonding is a weak chemical force that has so far mostly found applications in crystal engineering. Despite its potential for use in drug discovery, as a new molecular tool in the direction of molecular recognition events, it has rarely been assessed in biopolymers. Motivated by this fact, we have developed a peptide model system that permits the quantitative evaluation of weak forces in a biologically relevant proteinlike environment and have applied it for the assessment of a halogen bond formed between two amino acid side chains. The influence of a single weak force is measured by detection of the extent to which it modulates the conformation of a cooperatively folding system. We have optimized the amino acid sequence of the model peptide on analogues with a hydrogen bond-forming site as a model for the intramolecular halogen bond to be studied, demonstrating the ability of the technique to provide information about any type of weak secondary interaction. A combined solution nuclear magnetic res...
TL;DR: The novel class of Halo-tag fluorogenic probes that are derived from solvatochromic fluorophores enables a gel-free quantification of fusion proteins expressed in mammalian cells, an application that was not possible with previously nonfluorogenic Halo- tag probes.
Abstract: The design of fluorogenic probes for a Halo tag is highly desirable but challenging. Previous work achieved this goal by controlling the chemical switch of spirolactones upon the covalent conjugation between the Halo tag and probes or by incorporating a “channel dye” into the substrate binding tunnel of the Halo tag. In this work, we have developed a novel class of Halo-tag fluorogenic probes that are derived from solvatochromic fluorophores. The optimal probe, harboring a benzothiadiazole scaffold, exhibits a 1000-fold fluorescence enhancement upon reaction with the Halo tag. Structural, computational, and biochemical studies reveal that the benzene ring of a tryptophan residue engages in a cation−π interaction with the dimethylamino electron-donating group of the benzothiadiazole fluorophore in its excited state. We further demonstrate using noncanonical fluorinated tryptophan that the cation−π interaction directly contributes to the fluorogenicity of the benzothiadiazole fluorophore. Mechanistically, t...