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The glyoxalase pathway plays a vital role in the chemical detoxification of methylglyoxal (MG) in biological systems. Our previous study suggested that OsGLYI3 may be effective in seed natural aging. In this study, the rice OsGLYI3 gene was cloned and characterized as specifically expressed in the seed. The accelerated aging (AA) treatment results indicated significant roles of OsGLYI3 in seed longevity and vigor, as the seeds of the transgenic lines with overexpressed and knocked-out OsGLYI3 exhibited higher and lower germination, respectively. The AA treatment also increased the superoxide dismutase (SOD) activity in the overexpressed transgenic seeds compared to the wild-type seeds yet lowered the SOD activity in the CRISPR/Cas9-derived transgenic rice lines. Rice OsGLYI3 was markedly upregulated in response to NaCl induced stress conditions. Compared to wild-type plants, overexpressed transgenic rice lines exhibited increased GLYI activity, decreased MG levels and improved salt stress tolerance, while CRISPR/Cas9 knockout transgenic rice lines showed decreased glyoxalase I activity, increased MG levels, and greater sensitivity to stress treatments with NaCl. Collectively, our results confirmed for the first time that OsGLYI3 is specifically expressed in rice seeds and contributes to seed longevity and salt stress tolerance. • OsGLYI3 was confirmed for the first time specifically expressed in rice seed. • OsGLYI3 exhibits a detoxification function on methylglyoxal (MG). • OsGLYI3 plays an important role in rice seed longevity and vigor. • OsGLYI3 is essential for stress tolerance improvement in rice. [ABSTRACT FROM AUTHOR]

The accumulation of a metabolic by product – methylglyoxal above a minimal range can be highly toxic in all organisms. Stress induced elevation in methylglyoxal inactivates proteins and nucleic acids. Glutathione dependent glyoxalase enzymes like glyoxalase I and glyoxalase II together with glutathione independent glyoxalase III play inevitable role in methylglyoxal detoxification. Glyoxalase genes are generally conserved but with obvious exceptions. Mangroves being potent harsh land inhabitants, their internal organelles are constantly been exposed to elevated levels of methylglyoxal. First and foremost it is important to detect the presence of glyoxalases in mangroves. De novo transcriptome analysis of mangrove species Rhizophora mucronata Lam., identified eleven putative glyoxalase proteins (RmGLYI-1 to 5, RmGLYII-1 to 5 and RmGLYIII). Molecular characterization proposed PLN02300 or PLN02367 as the key domains of RmGLYI proteins. They possess molecular weight ranging from 26.45 to 32.53 kDa and may localize in cytosol or chloroplast. RmGLYII proteins of molecular weight 28.64–36 kDa, carrying PLN02398 or PLN02469 domains are expected to be localized in diverse cellular compartments. Cytosolic RmGLYIII with DJ-1/PfpI domain carries a molecular weight 26.4 kDa. Detailed structural analysis revealed monomeric nature of RmGLYI-1 and RmGLYII-1 whereas RmGLYIII is found to be homodimer. Molecular phylogenetic analysis and multiple sequence alignment specified conserved metal ion/substrate binding residues of RmGLY proteins. Estimation of relative expression of glyoxalases under salt stress indicated the prominence of RmGLYI and RmGLYII over RmGLYIII. The aforementioned prominence is supported by salt induced expression difference of glutathione metabolic enzymes and glutathione regulated transporter protein. • Reference sequence and annotation datasets from the de novo transcriptome of salt tolerant mangrove, Rhizophora mucronata. • GSH dependent and independent glyoxalase system in extremophilic mangroves. • Structural/biochemical characteristics of mangrove glyoxalase proteins. • Stability of glyoxalase structure models under salt stress; a lead for unidentified stability aspects of mangrove proteins. • Salt induced gene expression of mangrove glyoxalases correlated to GSH metabolic enzymes and GSH regulated transporter. [ABSTRACT FROM AUTHOR]

The detoxification of cadmium (Cd) or chromium (Cr) by microorganisms plays a vital role in bacterial survival and restoration of the polluted environment, but how microorganisms detoxify Cd and Cr simultaneously is largely unknown. Here, we isolated a bacterium, Cupriavidus sp. MP-37, which immobilized Cd(II) and reduced Cr(VI) simultaneously. Notably, strain MP-37 exhibited variable Cd(II) immobilization phenotypes, namely, cell adsorption and extracellular immobilization in the co-presence of Cd(II) and Cr(VI), while cell adsorption in the presence of Cd(II) alone. To unravel Cr(VI)-induced extracellular Cd(II) immobilization, proteomic analysis was performed, and methylglyoxal-scavenging protein (glyoxalase I, GlyI) and a regulator (YafY) showed the highest upregulation in the co-presence of Cd(II) and Cr(VI). GlyI overexpression reduced the intracellular methylglyoxal content and increased the immobilized Cd(II) content in extracellular secreta. The addition of lactate produced by GlyI protein with methylglyoxal as substrate increased the Cd(II) content in extracellular secreta. Reporter gene assay, electrophoretic mobility shift assay, and fluorescence quenching assay demonstrated that glyI expression was induced by Cr(VI) but not by Cd(II), and that YafY positively regulated glyI expression by binding Cr(VI). In the pot experiment, inoculation with the MP-37 strain reduced the Cd content of Oryza sativa L., and their secreted lactate reduced the Cr accumulation in Oryza sativa L. This study reveals that Cr(VI)-induced detoxification system drives methylglyoxal scavenging and Cd(II) extracellular detoxification in Cd(II) and Cr(VI) co-existence environment. [Display omitted] • Strain MP-37 displays simultaneous Cd(II) removal and Cr(VI) reduction abilities. • A Cr(VI)-induced methylglyoxal detoxification system increased Cd(II) removal. • Yafy and GlyI are key factors in the methylglyoxal detoxification system. • YafY positively regulated glyI expression, and GlyI was a Cr(VI)-binding protein. • The variable detoxification strategy of Cd(II) by bacteria favors survival. [ABSTRACT FROM AUTHOR]

• Biosensor for the detection of diabetes biomarker 'methylglyoxal' in blood plasma. • Sensitivity of 1.02 mA µM−1 and LOD of 0.21 µM were observed. • Biosensor responses for 350 human blood plasma samples were recorded. • Cross-validation of the biosensor with ELISA technique showed a 90% of correlation. Methylglyoxal (MG), a highly reactive by-product of glycolysis, is involved in the formation of advanced glycation end-products (AGEs). Elevated levels of MG have been correlated with micro-and macro-angiopathic complications in diabetes, including neuropathy, kidney disease, retinopathy, and cardiovascular disease. Therefore, point-of-care devices for detecting MG may be of great use in the screening of diabetes complications. This study was designed to determine the utility of the developed electrochemical biosensor to measure the level of MG in human plasma from type-2 diabetes mellitus patients. Electrochemical studies were carried out with optimized experimental parameters using the modified Platinum-electrode. Subsequently, clinical studies using 350 blood plasma samples were conducted and the results were validated against the ELISA kit, Normal Glucose Tolerance (NGT), and glycosylated haemoglobin (HbA1c). The MG sensor exhibited a linear range of 1.0–7.5 μM concentration with a sensitivity of 1.02 mA µM−1, a limit of detection of 0.21 µM, a limit of quantification of 0.70 µM and a response time less than 10 s. The sensor showed 90% correlation with ELISA data. The developed biosensor showed a significant correlation with HbA1c and fasting plasma glucose suggesting that it can be used as a point-of-care device to screen for diabetes. [ABSTRACT FROM AUTHOR]

Methylglyoxal is a common cytotoxic metabolite produced in plants during multiple biotic and abiotic stress. To mitigate the toxicity of MG, plants utilize the glyoxalase pathway comprising glyoxalase I (GLYI), glyoxalase II (GLYII), or glyoxalase III (GLYIII). GLYI and GLYII are the key enzymes of glyoxalase pathways that play an important role in abiotic stress tolerance. Earlier research showed that MG level is lower when both GLYI and GLYII are overexpressed together, compared to GLYI or GLYII single gene overexpressed transgenic plants. D-lactate dehydrogenase (D-LDH) is an integral part of MG detoxification which metabolizes the end product (D-lactate) of the glyoxalase pathway. In this study, two Arabidopsis transgenic lines were constructed using gene pyramiding technique: GLYI and GLYII overexpressed (G-I + II), and GLYI, GLYII, and D-LDH overexpressed (G-I + II + D) plants. G-I + II + D exhibits lower MG and D-lactate levels and enhanced abiotic stress tolerance than the G-I + II and wild-type plants. Further study explores the stress tolerance mechanism of G-I + II + D plants through the interplay of different regulators and plant hormones. This, in turn, modulates the expression of ABA-dependent stress-responsive genes like RAB18, RD22, and RD29B to generate adaptive responses during stress. Therefore, there might be a potential correlation between ABA and MG detoxification pathways. Furthermore, higher STY46, GPX3, and CAMTA1 transcripts were observed in G-I + II + D plants during abiotic stress. Thus, our findings suggest that G-I + II + D has significantly improved MG detoxification, reduced oxidative stress-induced damage, and provided a better protective mechanism against abiotic stresses than G-I + II or wild-type plants. [Display omitted] • Gene pyramiding was done to combine the Glyoxalase pathway along with D-LDH (G-I+II+D). • A comparative analysis was done between G-I+II+D with G-I+II (Glyoxalase I and II overexpressed) transgenics. • Pyramiding D-LDH with glyoxalase pathway (G-I+II+D) enhances abiotic stress tolerance in Arabidopsis thaliana. • G-I+II+D transgenics maintain lower methylglyoxal, D-Lactate, and ROS than G-I+II and wild-type plants during abiotic stress. [ABSTRACT FROM AUTHOR]

Objectives: Non-enzymatic glycation is closely associated with altered mechanical properties of connective tissue. Pregnancy, marked with high levels of female hormones, confers unique alteration to the mechanical properties of pelvic connective tissues in order to meet their physiological demands. However, there are few studies on glycation content and its influence on the mechanical properties of pelvic connective tissues during pregnancy. We hypothesise that the glycation content in pelvic tissues will change with a corresponding alteration in their mechanical properties, and that these changes are influenced by hormone levels. This study aims to investigate the correlation of vaginal tissue glycation content and mechanical property changes during pregnancy in association with the expression of a key pregnancy hormone (oestrogen) receptor, and an antioxidant enzyme, glyoxalase I.Study Design: A rat vaginal tissue model (tissues from non-pregnant and E15-E18 (last trimester) pregnant rats) was used in this study. Mechanical characteristics of vaginal tissues were analysed by a ball-indentation technique while modulus and morphology of the collagen fibrils within the tissues were measured with atomic force microscopy. A glycation marker, pentosidine, was quantified by a high performance liquid chromatography. The expression of oestrogen receptor and glyoxalase I in the tissue was qualified by immunochemical staining. The glycosaminoglycan (GAG) concentration difference in the tissues were quantified by a biochemical assay.Results: Pregnant rat vaginal tissue was characterised by significantly lower amounts of pentosidine, higher oestrogen receptor and glyoxalase I expression with larger creep, lower elastic modulus, larger fibril diameter and higher GAG content than their non-pregnant counterpart. There was a negative correlation between pentosidine and vaginal tissue creep.Conclusion: There was a reduction in vaginal tissue pentosidine in pregnancy with an associated increase in oestrogen receptor and glyoxalase I immunoexpression. Reduced glycation was associated with increased creeping of vaginal tissue. Oestrogen may therefore play a role in the increase of the vaginal wall's capacity to stretch through glyoxalase I up-regulation and subsequent glycation reduction. The new insight of the correlation of women's oestrogen level, glycation reaction and pelvic tissue mechanical property from this study may enhance our understanding of some pelvic organ diseases. [ABSTRACT FROM AUTHOR]

Human glyoxalase I (GLO I), a rate-limiting enzyme for detoxification of methylglyoxal (MG), a by-product of glycolysis, is known to be a potential therapeutic target for cancer. Here, we searched new scaffolds from natural compounds for designing novel GLO I inhibitors and found trans -stilbene scaffold. We examined the inhibitory abilities to human GLO I of commercially available trans- stilbene compounds. Among them, piceatannol was found to have the most potent inhibitory activity against human GLO I. Piceatannol could inhibit the proliferation of human lung cancer NCI-H522 cells, which are dependent on GLO I for survival, in a dose- and time-dependent manner. In addition, piceatannol more significantly inhibited the proliferation of NCI-H522 cells than that of NCI-H460 cells, which are less dependent on GLO I. Importantly, overexpression of GLO I in NCI-H522 cells resulted in less sensitive to the antiproliferative activity of piceatannol. Taken together, this is the first report demonstrating that piceatannol inhibits GLO I activity and the GLO I-dependent proliferation of cancer cells. Furthermore, we determined a pharmacophore for novel inhibitors of human GLO I by computational simulation analyses of the binding mode of piceatannol to the enzyme hot spot in the active site. We suggest that piceatannol is a possible lead compound for the development of novel GLO I inhibitory anticancer drugs. [ABSTRACT FROM AUTHOR]

Glyoxalase I (Gly I) is a component of the glyoxalase system which is involved in the detoxification of methylglyoxal, a byproduct of glycolysis. In the present study, a gene of rice ( Oryza sativa L., cv. Nipponbare) encoding Gly I was cloned and characterized. The quantitative real-time PCR analysis indicated that rice Gly I ( OsGly I ) was ubiquitously expressed in root, stem, leaf, leaf sheath and spikelet with varying abundance. OsGly I was markedly upregulated in response to NaCl, ZnCl 2 and mannitol in rice seedlings. For further functional investigation, OsGly I was overexpressed in rice using Agrobacterium- mediated transformation. Transgenic rice lines exhibited increased glyoxalase enzyme activity, decreased methylglyoxal level and improved tolerance to NaCl, ZnCl 2 and mannitol compared to wild-type plants. Enhancement of stress tolerance in transgenic lines was associated with reduction of malondialdehyde content which was derived from cellular lipid peroxidation. In addition, the OsGly I -overexpression transgenic plants performed higher seed setting rate and yield. Collectively, these results indicate the potential of bioengineering the Gly I gene in crops. [ABSTRACT FROM AUTHOR]

Density functional theory has been used to study the mechanism and stereospecificity of the catalytic reaction of human glyoxalase I. We used the quantum mechanical cluster method to model the enzyme active site. Glyoxalase I accepts both enantiomers of the hemithioacetal between methylglyoxal and glutathione and converts them to the S -D enantiomer of lactoylglutathione. We have compared several previously suggested or alternative reaction mechanisms for both substrates on an equal footing. The results show that the coordination shell of the Zn ion in the optimized geometries is more symmetric than in some inhibitor crystal structures, which we assign to differences in the electronic structure and the protonation states of the substrate. The symmetry of the active site model indicates that the enzyme can use the same reaction mechanism for the S and the R enantiomers of the substrate, but with exchanged roles of the two active-site glutamate residues. However, the calculations show some asymmetry (0–4 kcal mol −1 differences in reaction energies and activation barriers), caused by the different coordination states of the glutamate residues in the starting crystal structure. Our results indicate that the only possibility for the stereospecificity of glyoxalase I is differences in the electrostatic surroundings and flexibility of the glutamate residues in the active site owing to their neighboring residues in the protein. [ABSTRACT FROM AUTHOR]

Background Recent studies indicate that increased expression of glyoxalase I (GLO1) could result in epileptic seizures; thus, this study further explored the association of GLO1 with epilepsy from the perspective of molecular genetics. Material and methods GLO1 single nucleotide polymorphisms (SNPs; rs1130534, rs4746 and rs1049346) were investigated in cohort I (the initial samples: 249 cases and 289 controls). A replication study designed to confirm the positive findings in cohort I was performed in cohorts II (the additional samples: 130 cases and 191 controls) and I + II. Results In cohorts I, II and I + II, the CC genotype at rs1049346 T > C exerts a protective effect against both late-onset epilepsy (odds ratio [OR] = 2.437, p = 0.013; OR = 2.844, p = 0.008; OR = 2.645, p = 0.000, q = 0.003, respectively) and drug-resistant epilepsy (DRE) (OR = 2.985, p = 0.020; OR = 2.943, p = 0.014; OR = 3.049, p = 0.001, q = 0.006, respectively). Further analyses in cohort I + II indicate that the presence of the TAC/AAT haplotypes (rs1130534–rs4746–rs1049346) may be used as a marker of predisposition to/protection against DRE ( p = 0.002, q = 0.010; p = 0.000, q = 0.002, respectively). Conclusions This study is the first to demonstrate that the GLO1 SNPs are significantly associated with epilepsy. In particular, the rs1049346 T > C SNPs are potentially useful for risk assessment of late-onset epilepsy and DRE. [ABSTRACT FROM AUTHOR]

Glyoxalase I (Glo1) is the main scavenging enzyme of methylglyoxal (MG), a potent precursor of advanced glycation end products (AGEs). AGEs are known to control multiple biological processes, including epithelial to mesenchymal transition (EMT), a multistep phenomenon associated with cell transformation, playing a major role in a variety of diseases, including cancer. Crystalline silica is a well-known occupational health hazard, responsible for a great number of human pulmonary diseases, such as silicosis. There is still much debate concerning the carcinogenic role of crystalline silica, mainly due to the lack of a causal demonstration between silica exposure and carcinogenesis. It has been suggested that EMT might play a role in crystalline silica-induced lung neoplastic transformation. The aim of this study was to investigate whether, and by means of which mechanism, the antiglycation defence Glo1 is involved in Min-U-Sil 5 (MS5) crystalline silica-induced EMT in BEAS-2B human bronchial epithelial cells chronically exposed, and whether this is associated with the beginning of a neoplastic-like transformation process. By using gene silencing/overexpression and scavenging/inhibitory agents, we demonstrated that MS5 induced hydrogen peroxide-mediated c-Jun-dependent Glo1 up-regulation which resulted in a decrease in the Argpyrimidine-modified Hsp70 protein level which triggered EMT in a novel mechanism involving miR-21 and SMAD signalling. The observed EMT was associated with a neoplastic-like phenotype. The results obtained provide a causal in vitro demonstration of the MS5 pro-carcinogenic transforming role and more importantly they provide new insights into the mechanisms involved in this process, thus opening new paths in research concerning the in vivo study of the carcinogenic potential of crystalline silica. [ABSTRACT FROM AUTHOR]

Silver's antimicrobial properties have been known for centuries, but exactly how it kills bacteria is still a mystery. Information on the competition between the native Ni2+ and abiogenic Ag+ cations in bacterial systems is also critically lacking. For example, urease, a famous nickel-containing enzyme that hydrolyzes urea into carbon dioxide and ammonia (a key step in the biogeochemical nitrogen cycle on Earth), is inhibited by Ag+ cations, but the molecular mechanism of silver's action is poorly understood. By employing density functional theory (DFT) calculations combined with the polarizable continuum model (PCM) computations we assess the susceptibility of the mono/binuclear Ni2+ binding sites in the nickel enzymatic centers to Ni2+→Ag+ substitution. The active centers in the mononuclear glyoxalase I and acireductone dioxygenase enzymes appear to be well protected against Ag+ attack and, presumably, stay functional even in its presence. On the other hand, the binuclear nickel binding site in urease appears vulnerable to silver attack - the results obtained are in line with available experimental data and give reason to assume a possible substitution of the essential Ni2+ cation from the urease metal center by Ag+. [Display omitted] • Factors controlling Ag+/Ni2+ competition in bacterial metalloproteins are elucidated. • Mononuclear Ni2+ binding sites appear protected against the attacks from Ag+. • Charged or highly polarizable ligands boost the Ni2+/Ag+ competitiveness. • Reduced solvent exposure of the binding pocket is beneficial for Ni2+→Ag+ exchange. • Binuclear Ni2+ binding site in urease appears vulnerable to silver attack. [ABSTRACT FROM AUTHOR]

The zinc metalloenzyme glyoxalase I (GlxI) catalyzes the glutathione-dependent inactivation of cytotoxic methylglyoxal. Two competitive bivalent GlxI inhibitors, polyBHG2-62 ( K i = 1.0 nM) and polyBHG2-54 ( K i = 0.3 nM), were synthesized based on the transition-state analog S -( N -bromophenyl- N -hydroxycarbamoyl) glutathione (BHG). The most effective inhibitor, polyBHG2-54, is the first subnanomolar inhibitor of GlxI, and is over 50-fold more potent than BHG itself. [ABSTRACT FROM AUTHOR]

Glyoxalase I (Glo1) is a cellular defense enzyme involved in the detoxification of methylglyoxal (MG), a cytotoxic by-product of glycolysis, and MG-derived advanced glycation end products (AGEs). Argpyrimidine (AP), one of the major AGEs coming from MG modification of protein arginines, is a proapoptotic agent. Crystalline silica is a well-known occupational health hazard, responsible for a relevant number of pulmonary diseases. Exposure of cells to crystalline silica results in a number of complex biological responses, including apoptosis. The present study was aimed at investigating whether, and through which mechanism, Glo1 was involved in Min-U-Sil 5 crystalline silica-induced apoptosis. Apoptosis, by TdT-mediated dUTP nick-end labeling assay, and transcript and protein levels or enzymatic activity, by quantitative real-time PCR, Western blot, and spectrophotometric methods, respectively, were evaluated in human bronchial BEAS-2B cells exposed or not (control) to crystalline silica and also in experiments with appropriate inhibitors. Reactive oxygen species were evaluated by coumarin-7-boronic acid or Amplex red hydrogen peroxide/peroxidase methods for peroxynitrite (ONOO − ) or hydrogen peroxide (H 2 O 2 ) measurements, respectively. Our results showed that Min-U-Sil 5 crystalline silica induced a dramatic ONOO − -mediated inhibition of Glo1, leading to AP-modified Hsp70 protein accumulation that, in a mechanism involving JNK and NF-κB, triggered an apoptotic mitochondrial pathway. Inhibition of Glo1 occurred at both functional and transcriptional levels, the latter occurring via ERK1/2 MAPK and miRNA 101 involvement. Taken together, our data demonstrate that Glo1 is involved in the Min-U-Sil 5 crystalline silica-induced BEAS-2B cell mitochondrial apoptotic pathway via a novel mechanism involving Hsp70, JNK, and NF-κB. Because maintenance of an intact respiratory epithelium is a critically important determinant of normal respiratory function, the knowledge of the mechanisms underlying its disruption may provide insight into the genesis, and possibly the prevention, of a number of pathological conditions commonly occurring in silica dust occupational exposure. [ABSTRACT FROM AUTHOR]

Abstract: Methylglyoxal is one of the most powerful glycating agents of proteins and other important cellular components and has been shown to be toxic to cultured cells. Methylglyoxal cytotoxicity appears to occur through cell-cycle arrest but, more often, through induction of apoptosis. In this study we examined whether, and through which molecular mechanism, methylglyoxal affects the growth of poorly aggressive LNCaP and invasive PC3 human prostate cancer cells, where its role has not been exhaustively investigated yet. We demonstrated that methylglyoxal is cytotoxic on LNCaP and PC3 and that such cytotoxicity occurs not via cell proliferation but apoptosis control. Moreover, we demonstrated that methylglyoxal cytotoxicity, potentiated by the silencing of its major scavenging enzyme Glyoxalase I, occurred via different apoptotic responses in LNCaP and PC3 cells that also showed a different susceptibility to this metabolite. Finally, we showed that the observed methylglyoxal apoptogenic role involved different molecular pathways, specifically mediated by methylglyoxal or methylglyoxal-derived argpyrimidine intracellular accumulation and NF-kB signaling-pathway. In particular, in LNCaP cells, methylglyoxal, through the accumulation of argpyrimidine, desensitized the key cell survival NF-kB signaling pathway, which was consistent with the modulation of NF-kB-regulated genes, triggering a mitochondrial apoptotic pathway. The results suggest that this physiological compound merits investigation as a potential chemo-preventive/-therapeutic agent, in differently aggressive prostate cancers. [Copyright &y& Elsevier]

Abstract: Objectives: The aim of the study was to analyze polymorphisms of receptor for advanced glycation end products (RAGE) gene, and glyoxalase I gene and soluble RAGE, sRAGE, in physiological and pathological pregnancy. Design and methods: Polymorphisms of RAGE gene (−429 T/C, −374 T/A, 557 G/A, 2184 A/G) and glyoxalase I gene (A419C) and sRAGE serum levels were determined in 284 women with pathological and physiological pregnancy. Results: No differences in distribution of genotype and allelic frequencies of studied polymorphisms were found. GA genotype of RAGE 557 G/A polymorphism (known as Gly82Ser) is associated with lower sRAGE serum levels in healthy pregnant women compared to GG genotype (483±104 vs. 692±262pg/mL, p=0.008). sRAGE correlates negatively with ALT in patients with pregnancy intrahepatic cholestasis (r=−0.536, p=0.05). Conclusions: We did not show any association of RAGE and glyoxalase I gene polymorphisms with pathological pregnancy, however further studies are needed to confirm the results. [Copyright &y& Elsevier]

Abstract: The human glyoxalase I (hGLO I), which is a rate-limiting enzyme in the pathway for detoxification of apoptosis-inducible methylglyoxal (MG), has been expected as an attractive target for the development of new anti-cancer drugs. We have previously identified a natural compound myricetin as a substrate transition-state (Zn2+-bound MG-glutathione (GSH) hemithioacetal) mimetic inhibitor of hGLO I. Here, we constructed a hGLO I/inhibitor 4-point pharmacophore based on the binding mode of myricetin to hGLO I. Using this pharmacophore, in silico screening of chemical library was performed by docking study. Consequently, a new type of compound, which has a unique benzothiazole ring with a carboxyl group, named TLSC702, was found to inhibit hGLO I more effectively than S-p-bromobenzylglutathione (BBG), a well-known GSH analog inhibitor. The computational simulation of the binding mode indicates the contribution of Zn2+-chelating carboxyl group of TLSC702 to the hGLO I inhibitory activity. This implies an important scaffold-hopping of myricetin to TLSC702. Thus, TLSC702 may be a valuable seed compound for the generation of a new lead of anti-cancer pharmaceuticals targeting hGLO I. [Copyright &y& Elsevier]

Abstract: Several recent developments suggest that the human glyoxalase I (GLO I) is a potential target for anti-tumor drug development. In present study, a series of curcumin derivatives with high inhibitory activity against human GLO I were discovered. Inhibition constant (K i) values of compounds 8, 9, 10, 11 and 13 to GLO I are 4.600μM, 2.600μM, 3.200μM, 3.600μM and 3.600μM, respectively. To elucidate the structural features of potent inhibitors, docking-based three-dimensional structure–activity relationship (3D-QSAR) analyses were performed. Satisfactory agreement between experiment and theory suggests that comparative molecular similarity index analysis (CoMSIA) modeling exhibit much better correlation and predictive power. The cross-validated q 2 value is 0.638 while no-validation r 2 value is 0.930. Integrated with docking-based 3D-QSAR CoMSIA modeling, molecular surface property (electrostatic and steric) mapping and molecular dynamics simulation, a set of receptor-ligand binding models and bio-affinity predictive models for rational design of more potent inhibitors of GLO I are established. [Copyright &y& Elsevier]

Abstract: Glyoxalase I (GLO I) is the rate-limiting enzyme for detoxification of methylglyoxal (MG), a side-product of glycolysis, which is able to induce apoptosis. Since GLO I is known to be highly expressed in the most tumor cells and little in normal cells, inhibitors of this enzyme has been expected to be new anticancer drugs. Here, we examined the inhibitory abilities to the human GLO I of anthocyanidins, such as delphinidin, cyanidin and pelargonidin. Among them, delphinidin was found to have the most potent inhibitory effect on human GLO I. Also, only delphinidin-induced apoptosis in HL-60 cells in a dose- and time-dependent manner. Furthermore, we determined a pharmacophore for delphinidin binding to the human GLO I by computational simulation analyses of the binding modes of delphinidin, cyanidin and pelargonidin to the enzyme hot spot. These results suggest that delphinidin could be a useful lead compound for the development of novel GLO I inhibitory anticancer drugs. [ABSTRACT FROM AUTHOR]

Abstract: Glyoxalase I (GLOI) is a key metalloenzyme in glycolytic pathway by detoxifying reactive α-ketoaldehydes such as methylglyoxal. Recent studies demonstrate that the nature product curcumin is an efficient inhibitor of GLOI, but its binding mechanism towards GLOI is still unclear. In the present study, molecular docking and molecular dynamics (MD) simulations were performed to better understand the inhibitory mechanism of curcumin towards GLOI. The enol form of curcumin coordinates with the catalytic zinc ion of GLOI and forms a strong hydrogen bond with Glu 172, whereas its keto tautomer displays unfavorable electrostatic interactions with Glu 172 and Glu 99. The calculated binding free energies suggest that GLOI prefers the primary enol form (ΔG =−30.38kcal/mol) to the keto tautomer (ΔG =−24.16kcal/mol). The present work also reveals that bisdemethoxycurcumin binds to GLOI in a similar manner as curcumin and exhibits a slightly less negative predicted binding free energy, which is further validated by our comparative kinetics analysis (Ki=18.2 and 10.3μM for bisdemethoxycurcumin and curcumin, respectively). Results of the study can provide an insight into the development of novel and more effective GLOI inhibitors. [Copyright &y& Elsevier]

Abstract: Glyoxalase I (GLO I) is the rate-limiting enzyme for detoxification of methylglyoxal (MG), a side product of glycolysis, which is able to induce apoptosis. Since GLO I is known to be highly expressed in the most tumor cells and little in normal cells, specific inhibitors of this enzyme have been expected as effective anticancer drugs. The purpose of this study is a good construction of the human GLO I/inhibitor pharmacophore to obtain unique human GLO I inhibitory seed compounds for the development of useful anticancer drugs. Here, we selected natural flavonoid compounds that possess a plane configuration of cis C-4 ketone and C-5 hydroxy groups as the substrate (MG) transition-state mimetic structure. These compounds were examined the inhibitory abilities to human GLO I activity and analyzed their structure–activity relationships to determine an important pharmacophore of flavonoids for the human GLO I binding. Our results point to the contribution of hydroxy groups at the B ring of flavonoids to the effective inhibition of the human GLO I. Based on the binding mode of flavonoids, we constructed the human GLO I/inhibitor pharmacophore. This work delivers the first three-dimensional (3D) structural data and explains certain flavonoids interact specifically with the human GLO I. [Copyright &y& Elsevier]

Abstract: The glyoxalase (Glx) system is a critical detoxification enzyme system that is widely distributed in prokaryotic and eukaryotic organisms. Glyoxalase I (GlxI), the first enzyme in the system, is a divalent metal-ion dependent lyase (isomerizing), and its homologs have recently been categorized into two metal activation classes which are either Zn2+-dependent or non-Zn2+ dependent (Ni2+-/Co2+-activated). The latter class encompasses enzymes of predominantly bacterial origin. We have identified two genes in Pseudomonas aeruginosa PAO1 encoding glyoxalase I enzymes in addition to the gloA1 sequence recently reported and characterized. The gloA1 and gloA2 genes encode non-Zn2+ dependent glyoxalase I enzymes and the gloA3 gene remarkably encodes a Zn2+-dependent homolog. To our knowledge this is the first report of a eubacterial species with several GlxI encoding genes, and also of an organism possessing GlxI enzymes from both metal activation classes. [Copyright &y& Elsevier]

Abstract: The present investigation relates the changes in the activity of glyoxalase enzymes (glyoxalase I, G I, and glyoxalase II, G II) in certain tissues (foot, gills and digestive gland) of the bivalve mollusc Scapharca inaequivalvis after a two-week exposure to a sublethal concentration (1.6μM, 0.1ppm) of Cu++ and untreated control specimens in marine water. The trials also included determination of Cu content in the tissues and measurements of lactate dehydrogenase (LDH) activity, to estimate either possible Cu accumulation or alterations in glycolytic activity. Copper accumulation was present in all the studied organs, being 45-fold increased in the gills. G I, G II and LDH activities were unchanged in the digestive gland after Cu exposure. G I activity in foot and gills of Cu-exposed animals decreased, likely due to a drop in the glycolytic rate, as pointed out by the parallel decrease of LDH activity. After Cu exposure, G II activity was unchanged in the gills, maybe to secure enough levels of GSH; however, in the foot G II activity decreased, likely giving an increased intracellular amount of S-d-lactoylglutathione as energy supplier. [Copyright &y& Elsevier]

Abstract: The zinc metalloenzyme glyoxalase I (GLO1) is thought to play a role in anxiety disorders because a reduced brain expression of GLO1 has been associated with increased anxiety-behaviours in mice. Recently, a functional Ala111Glu polymorphism in GLO1 has been shown to result in a reduced enzyme activity. The present study tested the hypothesis that this common genetic variant could confer susceptibility to panic disorder using an Italian population sample of 162 panic disorder patients and 288 matched controls. Statistical analysis failed to show association with the overall diagnosis of the disease. However, a weak but significant association was demonstrated between this polymorphism and panic disorder without agoraphobia. While our data suggest that this polymorphism is unlikely to have a major function in the pathogenesis of panic disorder, it could play a role in the subgroup of patients without agoraphobic avoidance. [Copyright &y& Elsevier]

The present study regards possible changes in the activity of glyoxalase system enzymes (glyoxalase I, GI, and glyoxalase II, GII) in tissues (brain, liver and white muscle) of the mediterranean bony fish Sparus auratus after a 20 days exposure to sublethal concentrations (0.1 or 0.5 ppm) of Cu in the marine water and on control untreated animals. The experiments also included measurements of copper concentration in the tissues, as well as of lactate dehydrogenase (LDH) activity, to evaluate possible Cu accumulation and changes in glycolytic activity respectively. Cu accumulation only occurs in the liver. GI, GII and LDH activities kept unchanged in the brain after copper exposure. GI activity in liver and muscle of copper-exposed animals decreases probably for a slackening in the glycolytic rate, as suggested by the lowering of LDH activity. GII activity remains unchanged or increases (liver extract, 0.5 ppm of Cu), maybe to safeguard enough cellular levels of GSH. [Copyright &y& Elsevier]

[Display omitted] • The carboxyl group in the TLSC702 molecule is critical for binding to GLO I. • Lipophilicity of TLSC702 derivatives is correlated with antiproliferative effects. • Addition of hydrophobic groups at phenyl position increases apoptosis inducibility. Glyoxalase I (GLO I) is a known therapeutic target in cancer. Even though TLSC702, a GLO I inhibitor that we discovered, induces apoptosis in tumor cells, exceptionally higher doses are required compared with those needed to inhibit GLO I activity in vitro. In this work, structure–activity optimization studies were conducted on four sections of the TLSC702 molecule to determine the partial structural features necessary for the inhibition of GLO I. Herein, we found that the carboxy group in TLSC702 was critical for binding with the divalent zinc at the active site of GLO I. In contrast, the side chain substituents in the meta- and para- positions of the benzene ring had little influence on the in vitro inhibition of GLO I. The CLogP values of the TLSC702 derivatives showed a positive correlation with the antiproliferative effects on NCI-H522 cells. Thus, two derivatives of TLSC702, which displayed either high or low lipophilicity due to the types of substituents at the phenyl position, were selected. Even though both derivatives showed comparable inhibitory effects as that of their parent compound, the derivative with the high CLogP value was distinctly more antiproliferative than TLSC702. In contrast, the derivative with the low CLogP value did not decrease cell viability in NCI-H522 and HL-60 cells. These findings suggested that structural improvements, such as the addition of hydrophobic moieties to the phenyl group, enhanced the ability of TLSC702 to induce apoptosis by increasing cell membrane permeability. [ABSTRACT FROM AUTHOR]

• Four different QM-clusters were used to study the catalytic reaction of Glyoxalase I. • A QM-cluster with a small Glu-99 can produce the mechanism of the S substrate. • The same cluster predicts relative stabilities of the substrate's binding modes. • The same cluster cannot produce the mechanism of the R substrate. • It is better to use QM/MM methods to study reactions of stereospecific enzymes. Quantum mechanical (QM) cluster approach is a useful and convenient tool to study mechanisms of enzymatic reactions. Our investigation is to propose a proper QM-cluster that reproduces the special specificity of glyoxalase I (GlxI). GlxI converts S and R enantiomers of hemithioacetal to only S -D enantiomer of its product. It shows this special specificity with two glutamate residues that are symmetrically coordinated to a Zn ion. However, molecular dynamics simulations showed that one of the glutamates, Glu-172 is more flexible than the other one, Glu-99. Results indicate that a QM-cluster with a more flexible model of Glu-172 can reproduce proposed mechanisms for the S -substrate. However, the same cluster can reach to the R -L enantiomer of the product from the R substrate. We propose the necessity of using much more expensive, hybrid QM/MM methods or a very big QM-cluster to model and study reaction mechanisms of stereospecific enzymes like GlxI. [ABSTRACT FROM AUTHOR]

• Anabaena sp. PCC7120 genome possesses only Alr2321 as a Ni2+dependent active glyoxalase I enzyme like other prokaryotes. • Alr2321 is a key enzyme of Anabaena sp. PCC7120 for toxic methylglyoxal (MG) detoxification. • alr2321 transcript was induced under a range of abiotic stresses. • Alr2321 provides tolerance to cells against various abiotic stresses through detoxification of MG and reactive oxygen species. Abiotic stresses enhance the cellular level of reactive oxygen species (ROS) which consequently leads to toxic methylglyoxal (MG) production. Glyoxalases (GlyI & GlyII) catalyze the conversion of toxic MG into non-toxic lactic acid but their properties and functions have been overlooked in cyanobacteria. This is the first attempt to conduct a genome-wide analysis of GlyI protein (PF00903) from Anabaena sp. PCC7120. Out of total nine GlyI domain possessing proteins, only three (Alr2321, Alr4469, All1022) harbour conserve His/Glu/His/Glu metal binding site at their homologous position and are deficient in conserved region specific for Zn2+ dependent members. Their biochemical, structural and functional characterization revealed that only Alr2321 is a homodimeric Ni2+ dependent active GlyI with catalytic efficiency 11.7 × 106 M−1 s−1. It has also been found that Alr2321 is activated by various divalent metal ions and has maximum GlyI activity with Ni2+ followed by Co2+ > Mn2+ > Cu2+ and no activity with Zn2+. Moreover, the expression of alr2321 was found to be maximally up-regulated under heat (19 fold) followed by cadmium, desiccation, arsenic, salinity and UV-B stresses. BL21/pGEX-5X2- alr2321 showed improved growth under various abiotic stresses as compared to BL21/pGEX-5X2 by increased scavenging of intracellular MG and ROS levels. Taken together, these results suggest noteworthy links between intracellular MG and ROS, its detoxification by Alr2321, a member of GlyI family of Anabaena sp. PCC7120, in relation to abiotic stress. [ABSTRACT FROM AUTHOR]

Advanced glycation end products (AGEs) play an important role in the onset of diabetic retinopathy. Therefore, in the current study, we investigate whether and how Tanshinone IIa (Tan IIa) from Salvia miltiorrhiza protects bovine retinal endothelial cells (BRECs) against methylglyoxal (MGO) mediated cell dysfunction. The results showed that MGO reduced cell viability in dose dependent manner. The treatment of Tan IIa (50 μM) significantly improved cell viability induced by MGO in BRECs. MGO increased cellular reactive oxygen species formation and cellular nitric oxide (NO) level; enhanced nox1 and iNOS mRNA levels; inhibited prdx1 mRNA level. The treatment of Tan IIa effectually ameliorated cellular oxidative stress. Exposure of MGO resulted in mitochondrial fission and decrease of opa1 and mfn1. No significant difference in mRNA levels of mfn2 and drp1 was detected between MGO and medium. Tan IIa reduced mitochondrial fragmentation, enhanced the mRNA levels of mfn1 and opa1 in MGO cultured BRECs. The short time exposure of cellular antioxidatants, dimethylthiourea (10 mM) and tiron (10 mM) had no effect on mitochondrial fission although they ameliorated cellular reactive oxygen species level. Moreover, overexpression of glyoxalase 1 (GLO1) increased key proteins of mitochondrial fusion, including opa1 and mfn1 in BRECs cultured with MGO. However, inhibition of GLO1 by siRNA abolished the effect of Tan IIa on induction of mitochondrial fusion in MGO cultured BRECs. In conclusion, MGO caused the injury of retinal endothelial cells through induction of mitochondrial dysfunction and mitochondrial fission, the treatment of Tan IIa ameliorated mitochondrial dysfunction and fission induced by AGEs through enhancing GLO1. [ABSTRACT FROM AUTHOR]