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Cell Chemical Biology Jun 2017The guanidine-II (mini-ykkC) riboswitch is the smallest of the guanidine-responsive riboswitches, comprising two stem loops of similar sequence. We have solved...
The guanidine-II (mini-ykkC) riboswitch is the smallest of the guanidine-responsive riboswitches, comprising two stem loops of similar sequence. We have solved high-resolution crystal structures of both stem loops for the riboswitch from Gloeobacter violaceus. The stem loops have a strong propensity to dimerize by intimate loop-loop interaction. The dimerization creates specific binding pockets for two guanidine molecules, explaining their cooperative binding. Within the binding pockets the ligands are hydrogen bonded to a guanine at O6 and N7, and to successive backbone phosphates. Additionally they are each stacked upon a guanine nucleobase. One side of the pocket has an opening to the solvent, slightly lowering the specificity of ligand binding, and structures with bound methylguanidine, aminoguanidine, and agmatine show how this is possible.
Topics: Base Sequence; Conserved Sequence; Dimerization; Guanidine; Inverted Repeat Sequences; Ligands; Models, Molecular; Nucleic Acid Conformation; Riboswitch; Substrate Specificity
PubMed: 28529131
DOI: 10.1016/j.chembiol.2017.05.014 -
Macromolecular Bioscience Apr 2018Poly(N-methylvinylamines) with secondary amines can form complexes with plasmid DNA (pDNA) and provide transfection efficiency in HeLa cells in the same order as linear...
Poly(N-methylvinylamines) with secondary amines can form complexes with plasmid DNA (pDNA) and provide transfection efficiency in HeLa cells in the same order as linear polyethyleneimine but with higher cell viability. Chemical modifications of poly(N-methylvinylamine) backbones are performed to further improve transfection efficiency while maintaining low degree of cytotoxicity. In a first type of polymer, primary amino groups are incorporated via a copolymerization strategy. In a second one, primary amino and imidazole groups are incorporated also via a copolymerization strategy. In a third one, secondary amino groups are substituted with methylguanidine functions through a postpolymerization reaction. Thus, novel polymers of various molecular masses are synthesized, characterized, and their interaction with pDNA studied. Then, their transfection efficiency and cytotoxicity are tested in HeLa cells. Two polymethylvinylamine-based copolymers, one containing 20% of imidazole moieties and another one composed of 12% of guanidinyl units allow remarkable transfection efficiency of HeLa, pulmonary (16HBE), skeletal muscle (C2C12), and dendritic (DC2.4) cells. Overall, this work thus identifies new promising DNA carriers and chemical modifications that improve the transfection efficiency while maintaining low degree of cytotoxicity.
Topics: Cell Survival; DNA; Gene Transfer Techniques; HeLa Cells; Humans; Plasmids; Polymers; Transfection; Vinyl Compounds
PubMed: 29392880
DOI: 10.1002/mabi.201700353 -
Cell Chemical Biology Nov 2017Riboswitches are structural elements found in mRNA molecules that couple small-molecule binding to regulation of gene expression, usually by controlling transcription or...
Riboswitches are structural elements found in mRNA molecules that couple small-molecule binding to regulation of gene expression, usually by controlling transcription or translation. We have determined high-resolution crystal structures of the ykkC guanidine III riboswitch from Thermobifida fusca. The riboswitch forms a classic H-type pseudoknot that includes a triple helix that is continuous with a central core of conserved nucleotides. These form a left-handed helical ramp of inter-nucleotide interactions, generating the guanidinium cation binding site. The ligand is hydrogen bonded to the Hoogsteen edges of two guanine bases. The binding pocket has a side opening that can accommodate a small side chain, shown by structures with bound methylguanidine, aminoguanidine, ethylguanidine, and agmatine. Comparison of the new structure with those of the guanidine I and II riboswitches reveals that evolution generated three different structural solutions for guanidine binding and subsequent gene regulation, although with some common elements.
Topics: Actinobacteria; Base Sequence; Binding Sites; Crystallography, X-Ray; Guanidine; Guanine; Hydrogen Bonding; Models, Molecular; Nucleic Acid Conformation; Riboswitch
PubMed: 28988949
DOI: 10.1016/j.chembiol.2017.08.021 -
Frontiers in Public Health 2022Gemcitabine and cisplatin serve as appropriate treatments for patients with cholangiocarcinoma (CCA). Our previous study using histoculture drug response assay (HDRA),...
Gemcitabine and cisplatin serve as appropriate treatments for patients with cholangiocarcinoma (CCA). Our previous study using histoculture drug response assay (HDRA), demonstrated individual response patterns to gemcitabine and cisplatin. The current study aimed to identify predictive biomarkers for gemcitabine and cisplatin sensitivity in tissues and sera from patients with CCA using metabolomics. Metabolic signatures of patients with CCA were correlated with their HDRA response patterns. The tissue metabolic signatures of patients with CCA revealed the inversion of the TCA cycle that is evident with increased levels of citrate and amino acid backbones as TCA cycle intermediates, and glucose which corresponds to cancer stem cell (CSC) properties. The protein expression levels of CSC markers were examined on tissues and showed the significantly inverse association with the responses of patients to cisplatin. Moreover, the elevation of ethanol level was observed in gemcitabine- and cisplatin-sensitive group. In serum, a lower level of glucose but a higher level of methylguanidine was observed in the gemcitabine-responders as non-invasive predictive biomarker for gemcitabine sensitivity. Collectively, our findings indicate that these metabolites may serve as the predictive biomarkers in clinical practice which not only predict the chemotherapy response in patients with CCA but also minimize the adverse effect from chemotherapy.
Topics: Bile Duct Neoplasms; Bile Ducts, Intrahepatic; Biomarkers; Cholangiocarcinoma; Cisplatin; Deoxycytidine; Glucose; Humans; Gemcitabine
PubMed: 35223723
DOI: 10.3389/fpubh.2022.766023 -
Neuroscience Sep 2015Patients with chronic renal failure often have hypertension, but the cause of hypertension, other than an excess of body fluid, is not well known. We hypothesized that...
Patients with chronic renal failure often have hypertension, but the cause of hypertension, other than an excess of body fluid, is not well known. We hypothesized that the bulbospinal neurons in the rostral ventrolateral medulla (RVLM) are stimulated by uremic toxins in patients with chronic renal failure. To investigate whether RVLM neurons are sensitive to uremic toxins, such as uric acid, indoxyl sulfate, or methylguanidine, we examined changes in the membrane potentials (MPs) of bulbospinal RVLM neurons of Wister rats using the whole-cell patch-clamp technique during superfusion with these toxins. A brainstem-spinal cord preparation that preserved the sympathetic nervous system was used for the experiments. During uric acid, indoxyl sulfate, or methylguanidine superfusion, almost all the RVLM neurons were depolarized. To examine the transporters for these toxins on RVLM neurons, histological examinations were performed. The uric acid-, indoxyl sulfate-, and methylguanidine-depolarized RVLM neurons showed the presence of urate transporter 1 (URAT 1), organic anion transporter (OAT)1 or OAT3, and organic cation transporter (OCT)3, respectively. Furthermore, the toxin-induced activities of the RVLM neurons were suppressed by the addition of an anti-oxidation drug (VAS2870, an NAD(P)H oxidase inhibitor), and a histological examination revealed the presence of NAD(P)H oxidase (nox)2 and nox4 in these RVLM neurons. The present results show that uric acid, indoxyl sulfate, and methylguanidine directly stimulate bulbospinal RVLM neurons via specific transporters on these neurons and by producing oxidative stress. These uremic toxins may cause hypertension by activating RVLM neurons.
Topics: Animals; Anion Transport Proteins; Benzoxazoles; Enzyme Inhibitors; Indican; Medulla Oblongata; Membrane Glycoproteins; Membrane Potentials; Methylguanidine; NADPH Oxidase 2; NADPH Oxidase 4; NADPH Oxidases; Neurons; Neuroprotective Agents; Neurotoxins; Organic Anion Transport Protein 1; Organic Anion Transporters, Sodium-Independent; Oxidative Stress; Patch-Clamp Techniques; Rats, Wistar; Renal Insufficiency, Chronic; Sympathetic Nervous System; Triazoles; Uric Acid
PubMed: 26208844
DOI: 10.1016/j.neuroscience.2015.07.055 -
BMC Nephrology Apr 2023Chronic kidney disease (CKD) is a global public health issue. The diagnosis of CKD would be considerably enhanced by discovering novel biomarkers used to determine the...
BACKGROUND
Chronic kidney disease (CKD) is a global public health issue. The diagnosis of CKD would be considerably enhanced by discovering novel biomarkers used to determine the glomerular filtration rate (GFR). Small molecule metabolites related to kidney filtration function that might be utilized as biomarkers to measure GFR more accurately could be found via a metabolomics analysis of blood samples taken from individuals with varied glomerular filtration rates.
METHODS
An untargeted metabolomics study of 145 plasma samples was performed using ultrahigh-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS). The 145 samples were divided into four groups based on the patient's measured glomerular filtration rates (mGFRs) determined by the iohexol plasma clearance rate. The data were analyzed using random forest analyses and six other unique statistical analyses. Principal component analysis (PCA) was conducted using R software.
RESULTS
A large number of metabolites involved in various metabolic pathways changed significantly between groups with different GFRs. These included metabolites involved in tryptophan or pyrimidine metabolism. The top 30 metabolites that best distinguished between the four groups in a random forest plot analysis included 13 amino acids, 9 nucleotides, and 3 carbohydrates. A panel of metabolites (including hydroxyaparagine, pseudouridine, C-glycosyltryptophan, erythronate, N-acetylalanine, and 7-methylguanidine) for estimating GFR was selected for future testing in targeted analyses by combining the candidate lists with the six other statistical analyses. Both hydroxyasparagine and N,N-dimethyl-proline-proline are unique biomarkers shown to be inversely associated with kidney function that have not been reported previously. In contrast, 1,5-anhydroglucitol (1,5-AG) decreases with impaired renal function.
CONCLUSIONS
This global untargeted metabolomics study of plasma samples from patients with different degrees of renal function identified potential metabolite biomarkers related to kidney filtration. These novel potential metabolites provide more insight into the underlying pathophysiologic processes that may contribute to the progression of CKD, lead to improvements in the estimation of GFR and provide potential therapeutic targets to improve kidney function.
Topics: Humans; Glomerular Filtration Rate; Chromatography, Liquid; Tandem Mass Spectrometry; Renal Insufficiency, Chronic; Biomarkers
PubMed: 37085754
DOI: 10.1186/s12882-023-03147-9 -
Journal of Proteome Research Sep 2018Ureteral obstruction will lead clinically to hydronephrosis, which may further develop into partial or complete loss of kidney function and even cause permanent...
Ureteral obstruction will lead clinically to hydronephrosis, which may further develop into partial or complete loss of kidney function and even cause permanent histological damage. However, there is little knowledge of metabolic responses during the obstructed process and its recoverability. In this study, a complete unilateral ureteral obstruction (CUUO) model was established in the rabbit, and H NMR-based metabolomic analysis of urine was used to reveal the metabolic perturbations in rabbits caused by CUUO and the metabolic recovery after the CUUO was relieved. Univariate and multivariate statistical analyses were used to identify metabolic characteristics. The gradually decreased levels of 3-hydroxykynurenine, 3-methylhistidine, creatinine, guanidoacetate, meta- and para-hydroxyphenylacetate, and phenylacetylglycine and the gradually increased levels of acetate, alanine, citrate, glycine, lactate, and methionine in urine could be regarded as potential biomarkers for the occurrence and severity of ureteral obstruction. And the reduced levels of 3-methylhistidine, creatinine, guanidoacetate, hippurate, meta-hydroxyphenylacetate, and methylguanidine and the elevated levels of 2-aminoisobutyrate, acetylcholine, citrate, lactate, lysine, valine, and α-ketoglutarate in urine compared with the obstructed level could characterize the metabolic recovery of ureteral obstruction. Our results depicted the disturbed biochemical pathways involved in ureteral obstruction and demonstrated the practicability of recovering renal functions for the patients with severe hydronephrosis in clinical practice by removing causes for obstruction.
Topics: Acetic Acid; Alanine; Analysis of Variance; Animals; Biomarkers; Citric Acid; Creatinine; Disease Models, Animal; Glycine; Hydronephrosis; Kynurenine; Lactic Acid; Magnetic Resonance Spectroscopy; Male; Metabolome; Methionine; Methylhistidines; Phenylacetates; Rabbits; Ureter; Ureteral Obstruction
PubMed: 30024170
DOI: 10.1021/acs.jproteome.8b00304 -
Journal of Proteome Research Feb 2015Patients with pancreatic cancer (PC) are usually diagnosed at late stages, when the disease is nearly incurable. Sensitive and specific markers are critical for...
Patients with pancreatic cancer (PC) are usually diagnosed at late stages, when the disease is nearly incurable. Sensitive and specific markers are critical for supporting diagnostic and therapeutic strategies. The aim of this study was to use a metabonomics approach to identify potential plasma biomarkers that can be further developed for early detection of PC. In this study, plasma metabolites of newly diagnosed PC patients (n = 100) and age- and gender-matched controls (n = 100) from Connecticut (CT), USA, and the same number of cases and controls from Shanghai (SH), China, were profiled using combined gas and liquid chromatography mass spectrometry. The metabolites consistently expressed in both CT and SH samples were used to identify potential markers, and the diagnostic performance of the candidate markers was tested in two sample sets. A diagnostic model was constructed using a panel of five metabolites including glutamate, choline, 1,5-anhydro-d-glucitol, betaine, and methylguanidine, which robustly distinguished PC patients in CT from controls with high sensitivity (97.7%) and specificity (83.1%) (area under the receiver operating characteristic curve [AUC] = 0.943, 95% confidence interval [CI] = 0.908-0.977). This panel of metabolites was then tested with the SH data set, yielding satisfactory accuracy (AUC = 0.835; 95% CI = 0.777-0.893), with a sensitivity of 77.4% and specificity of 75.8%. This model achieved a sensitivity of 84.8% in the PC patients at stages 0, 1, and 2 in CT and 77.4% in the PC patients at stages 1 and 2 in SH. Plasma metabolic signatures show promise as biomarkers for early detection of PC.
Topics: Adult; Aged; Aged, 80 and over; Biomarkers; Chromatography, Gas; Chromatography, Liquid; Female; Humans; Male; Mass Spectrometry; Middle Aged; Pancreatic Neoplasms
PubMed: 25429707
DOI: 10.1021/pr501135f -
Neuropharmacology Dec 2019Poly(ADP-ribose) polymerase-1 (PARP1) is a ubiquitous nuclear enzyme that regulates DNA repair and genomic stability. In oxidative genotoxic conditions, PARP1 activity...
Poly(ADP-ribose) polymerase-1 (PARP1) is a ubiquitous nuclear enzyme that regulates DNA repair and genomic stability. In oxidative genotoxic conditions, PARP1 activity is enhanced significantly, leading to excessive depletion of nicotinamide adenine dinucleotide (NAD) and mitochondrial dysfunction. We hypothesized that PARP1-induced NAD depletion inhibits NAD-dependent sirtuin deacetylase activity, thereby interfering with the mitochondrial regulator, peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α). The DNA alkylator, N'-Nitro-N-nitroso-N-methylguanidine (MNNG), induced NAD depletion, inhibited sirtuin deacetylase activity and enhanced acetylation of PGC-1α. This was associated with reduced interaction between PGC-1α and nuclear respiratory factor 1 (NRF-1), which is a nuclear transcription factor that drives mitochondrial replication by regulating mitochondrial transcription factor A (TFAM). MNNG also reduced binding of NRF-1 to the tfam upstream promoter region and reduced TFAM mRNA, mitochondrial DNA copy number and respiratory function. MNNG effects were mitigated by PARP1 inhibition and genetic loss of function, by enhancing intracellular NAD levels, and with sirtuin (SIRT1) gain of function, supporting a mechanism dependent on PARP1 activity, NAD-depletion and SIRT1 inhibition. This and other work from our group supports a destructive sequelae of events related to PARP1-induced sirtuin inhibition and sirtuin-mediated regulation of transcription.
Topics: Acetylation; Animals; Cell Respiration; DNA, Mitochondrial; DNA-Binding Proteins; High Mobility Group Proteins; Methylnitronitrosoguanidine; Mice; Mitochondria; NAD; Neurons; Nuclear Respiratory Factor 1; Peroxisome Proliferator-Activated Receptor Gamma Coactivator 1-alpha; Poly (ADP-Ribose) Polymerase-1; Sirtuin 1; Transcription Factors; Transcription, Genetic
PubMed: 31487495
DOI: 10.1016/j.neuropharm.2019.107755 -
ACS Infectious Diseases Dec 2018Thirty-two new naphthylthiazole derivatives were synthesized with the aim of exploring their antimicrobial effect on multidrug-resistant Gram-positive bacteria....
Thirty-two new naphthylthiazole derivatives were synthesized with the aim of exploring their antimicrobial effect on multidrug-resistant Gram-positive bacteria. Compounds 25 and 32, with ethylenediamine and methylguanidine side chains, represent the most promising derivatives, as their antibacterial spectrum includes activity against multidrug-resistant staphylococcal and enterococcal strains. Moreover, the new derivatives are highly advantageous over the existing frontline therapeutics for the treatment of multidrug-resistant Gram-positive bacteria. In this vein, compound 25 possesses three attributes: no bacterial resistance was developed against it even after 15 passages, it was very efficient in targeting intracellular pathogens, and it exhibited a concentration-dependent ability to disrupt the preformed bacterial biofilm.
Topics: Animals; Anti-Bacterial Agents; Biofilms; Cell Line; Drug Design; Drug Resistance, Multiple, Bacterial; Enterococcus; Humans; Microbial Sensitivity Tests; Staphylococcal Infections; Staphylococcus aureus; Structure-Activity Relationship; Thiazoles
PubMed: 30247876
DOI: 10.1021/acsinfecdis.8b00172