Comparative Biochemical Analysis Research Articles

"Post-COVID-19 Vaccination: Comparative Biochemical Analysis In Normal Control, Type 2 Diabetic, And Hypertensive Individuals"

"Post-COVID-19 Vaccination: Comparative Biochemical Analysis In Normal Control, Type 2 Diabetic, And Hypertensive Individuals"

Comparative biochemical analysis of platelet-rich plasma-chitosan and platelet-rich fibrin-chitosan for treating tibial bone defects in rabbits.

Fracture healing can cause serious complications, both preoperatively and postoperatively, including malunion or non-union. Biomaterials can enhance the fracture healing process. This study aimed to compare platelet-rich plasma (PRP)-chitosan and platelet-rich fibrin (PRF)-chitosan on the basis of biochemical parameters for fracture treatment in rabbits. This study involved 12 clinically healthy rabbits. After preparing PRP and PRF, a 3-mm bone defect was created in the tibia of each rabbit. The animals were divided randomly into two groups (A and B). Group A received PRP-Chitosan, and Group B received PRF-Chitosan. Bone healing was assessed using biochemical parameters (calcium [Ca], phosphorus [P], serum alkaline phosphatase [ALP], and osteocalcin [Ocn]) at 2-, 4-, 6-, and 8-week postoperatively. The data were compared using repeated-measures analysis of variance (p < 0.05) with Statistical Package for the Social Sciences statistical software. Group-wise comparison showed no significant differences (p > 0.05) between the groups, except for ALP levels, which were significantly higher in Group B than in Group A (p < 0.05). In the week-wise comparison, there was a significant difference between both groups, as Ca and ALP levels showed significant differences at all weeks postoperatively, whereas Ocn showed a significant difference at 2- and 4-week postoperatively (p < 0.05). However, there was no difference in P levels between the groups at any post-operative week (p > 0.05). Both combinations enhanced bone regeneration. However, PRF-Chitosan is a better combination for bone repair than PRP-Chitosan. There were some limitations of this study, such as a small sample size, only male rabbits were used, and a lack of mechanical testing; these limitations should be addressed in future studies. The insights gained from the present study may open a new approach to the use of a combination of biomaterials for bone healing, which should be further investigated clinically and in other animal models as a future scope.

Comparative biochemical analysis of full-length and truncated BaqA α-amylases from Bacillus aquimaris MKSC 6.2

BaqA is a raw starch degrading α-amylase produced by the marine bacterium Bacillus aquimaris MKSC 6.2, associated with soft corals. This α-amylase belongs to a new subfamily Glycoside Hydrolases (GH) 13_45 which has several unique characteristics, namely, a pair of tryptophan residues Trp201 and Trp202, a distinct LPDIx signature in the Conserved Sequence Region-V (CSR-V), and an elongated C-terminus containing five aromatic residues. The research aims to investigate physicochemical, kinetics, and biochemical properties of BaqA. In this study, the full-length enzyme (BaqA) and a truncated form of BaqA (designated as BaqAΔC), lacking the C-terminal 34 amino acids were constructed and expressed in Escherichia coli ArcticExpress (DE3). BaqA formed inclusion bodies, while BaqAΔC was produced as a soluble protein. Purified and refolded BaqA exhibited a catalytic efficiency (kcat/Km) of 53.1 ± 6.3 mL mg−1 s−1 at 40 °C and pH 7.5, whereas the purified BaqAΔC displayed kcat/Km of 11.4 ± 1.3 mL mg−1 s−1 under the optimum condition of 50 °C and pH 6.5. Moreover, BaqAΔC showed a slight reduction in the binding affinity towards sago granules. Interestingly, BaqAΔC displayed robust stability and halotolerant properties compared to BaqA. BaqAΔC maintained 50 % amylolytic activity for up to 6 h, whereas BaqA lost over 50 % of its activity within 90 min. Furthermore, BaqAΔC showed a remarkable increase in amylolytic activity upon the addition of NaCl, with an optimum concentration of 0.5 M. Even at a high salt concentration (1.5 M NaCl), BaqAΔC retained over 50 % of its residual activity. Taken together, its solubility, amylolytic activity, stability, ability to degrade raw starch, and moderate halotolerance make BaqAΔC a promising candidate for various starch processing industries.

Exploring the Biochemical Profiles of Medicinal Plants Cultivated Under Stressful Environmental Conditions

The therapeutic potential of medicinal plants, attributed to diverse bioactive chemicals, is impacted by dynamic metabolic changes under environmental stressors such as salt, drought, and heavy metal exposure. This study aimed to elucidate the biochemical alterations in stressed medicinal plants and their implications for therapeutic qualities. Two groups of plants were compared: one cultivated in optimal conditions, serving as a control, and the other exposed to stressors mimicking real-world scenarios. Biochemical parameters, including total phenolic content, flavonoids, antioxidant activity, and specific secondary metabolites, were assessed using established analytical methods. In a case study involving Aloe vera under elevated soil salinity, a significant decrease in total phenolic content and antioxidant activity was observed. Surprisingly, aloin, an anti-inflammatory bioactive compound, exhibited notable upregulation. Stress-induced biochemical variations in medicinal plants differed significantly from those in ideal conditions, demonstrating adaptive responses, including modulation of secondary metabolite synthesis. While certain stresses led to an overall decrease in secondary metabolites, specific instances showed an overexpression of beneficial chemicals. Exploring drought stress in Lavandula angustifolia (lavender), Vitis vinifera, and Artemisia tridentata, revealed a marked reduction in flavonoids and specific secondary metabolites. The study aimed to establish connections between metabolic alterations and stress-adaptive reactions. The findings contribute to understanding how environmental stress influences the therapeutic efficacy of medicinal plants and shed light on the molecular pathways governing stress-induced changes. In the investigation of heavy metal exposure, Hypericum perforatum, and Coriandrum sativum subjected to cadmium-contaminated soil exhibited an overall decline in secondary metabolites, including reduced hypericin content. Contrarily, hyperforin, linked to antidepressant properties, experienced a significant increase. These results have implications for pharmaceutical applications, sustainable farming practices, and the conservation of medicinal plant biodiversity amidst changing environmental dynamics. The study underscores the intricate relationship between environmental stressors and the biochemical makeup of medicinal plants, emphasizing their resilience and capacity to produce valuable bioactive molecules in challenging conditions. The comparative biochemical analysis of stressed medicinal plants involves assessing various parameters to comprehend how stress influences plant metabolism and bioactive compound production.

Metabolic diversity, biosynthetic pathways, and metabolite biomarkers analysed via untargeted metabolomics and the antioxidant potential reveal for high temperature tolerance in tomato hybrid

High-temperature tolerant varieties/hybrids of tomato overcome stress-related changes due to their intrinsic cellular metabolic capabilities. We have used untargeted metabolite profiling and comparative biochemical and biological analyses of high temperature tolerant (HTT) tomato hybrid VRNTH18283 and sensitive (HTS) VRNTH19072 to decipher altered metabolic pathways and biomarker metabolites which were linked with enhanced antioxidant activity in HTT fruits. Parameters like fruit weight, yield per plant and key biochemical determinants viz. total soluble sugar (TSS), titratable acidity, ascorbic acid and lycopene content were significantly high in HTT hybrid compared to HTS. Metabolomics revealed upregulated metabolite diversity in HTT fruit extract (11,453 m/z features) as compared to 8834 m/z features in HTS. Statistical analysis revealed 423 differentially up-regulated and 410 down-regulated metabolite features. Multivariate data analyses showed sample discrimination based on distinct phytochemical diversification. Functionally annotated metabolite features showed enrichment in metabolic pathways including linoleic and linolenic acid metabolism, monoterpenoids biosynthesis and degradation, cutin, suberin and wax biosynthesis, and sphingolipids metabolism. Metabolite features were categorized in benzamides, amino acids, sugars, nucleotides, sterols, glycosides, phenylacetaldehydes, indoles, tryptamines and organic acids as major compound classes. Among 22 potentially discriminatory metabolites obtained from OPLS-DA model, all including prominently abundant solasodine, thiamine diphosphate, L-tryptophan, folic acid, lycopene, dihydrozeatin, myricetin, tomatidine, chlorogenic acid and α-tocotrienol were classified as metabolite biomarkers and were correlated with enhanced antioxidant activity. The study revealed that the altered physicochemical profile, increased metabolite compositional diversity and discriminatory metabolite biomarkers are liable for improved performance of the hybrid HTT VRNTH18283 against the negative impact of high temperature in comparison to HTS VRNTH19072.

A Comparative Biochemical and Pathological Evaluation of Brain Samples from Knock-In Murine Models of Gaucher Disease.

Gaucher disease (GD) is a lysosomal storage disorder stemming from biallelic mutations in GBA1, characterized by glucocerebrosidase dysfunction and glucocerebroside and glucosylsphingosine accumulation. Since phenotypes of murine models of GD often differ from those in patients, the careful characterization of Gba1 mutant mice is necessary to establish their ability to model GD. We performed side-by-side comparative biochemical and pathologic analyses of four murine Gba1 models with genotypes L444P/L444P (p.L483P/p.L483P), L444P/null, D409H/D409H (p.D448H/p.D448H) and D409H/null, along with matched wildtype mice, all with the same genetic background and cage conditions. All mutant mice exhibited significantly lower glucocerebrosidase activity (p < 0.0001) and higher glucosylsphingosine levels than wildtype, with the lowest glucocerebrosidase and the highest glucosylsphingosine levels in mice carrying a null allele. Although glucocerebrosidase activity in L444P and D409H mice was similar, D409H mice showed more lipid accumulation. No Gaucher or storage-like cells were detected in any of the Gba1 mutant mice. Quantification of neuroinflammation, dopaminergic neuronal loss, alpha-synuclein levels and motor behavior revealed no significant findings, even in aged animals. Thus, while the models may have utility for testing the effect of different therapies on enzymatic activity, they did not recapitulate the pathological phenotype of patients with GD, and better models are needed.

Biochemical Analysis and DNA Barcoding of Millet Echinochloa frumentacea

Background: Millets are small grains that are rich in nutrients. In recent times, millet-based foods have been increasingly recommended for a healthy diet. Many millets are not annotated or DNA barcoded yet. Methods: In this study, comparative biochemical analyses especially that of starch and total protein of Echinochloa frumentacea, called as Indian barnyard white millet (Varai), from geographically different locations like Tamil Nadu and Maharashtra have been done. Their DNA barcoding has also been done to identify them on the basis of molecular data. Result: It was observed that starch granules were more abundant in Tamil Nadu variety as compared to Maharashtra variety. Blue value, indicative of amylose: amylopectin ratio was found to be low in Varai, indicating that Varai has low starch digestibility and its starch releases glucose slowly, thus making it a low glycaemic index food. Protein content was higher in Tamil Nadu variety, but overall Varai had a lower protein content as compared to other millets. Ribulose-1,5-bisphosphate carboxylase/oxygenase large subunit (rbcL) gene from plastid was isolated, amplified by PCR, sequenced and the sequence was submitted to GenBank, NCBI. The gene was identified to be that of Echinochloa frumentacea and was given the accession numbers by GenBank as OR027010 (Varai, Maharashtra) and OR027011 (Varai, Tamil Nadu). This study indicated a distinct biochemical difference related to the geographical location of millets. This study helped barcoding of Echinochloa frumentacea Indian varieties using rbcL gene. This will further help in studies of phylogeny and evolution and also that of the relatedness of Echinochloa sp within and as compared to other millets.

Comparative biochemical analysis and GC–MS phytochemical profiling in some aquatic plants

Comparative biochemical analysis and GC–MS phytochemical profiling in some aquatic plants

AcHZP45 is a repressor of chlorophyll biosynthesis and activator of chlorophyll degradation in kiwifruit.

The degradation of chlorophyll during fruit development is essential to reveal a more 'ripe' color that signals readiness to wild dispersers of seeds and the human consumer. Here, comparative biochemical analysis of developing fruit of Actinidia deliciosa cv. Xuxiang ('XX', green-fleshed) and Actinidia chinensis cv. Jinshi No.1 ('JS', yellow-fleshed) indicated that variation in chlorophyll content is the major contributor to differences in flesh color. Four differentially expressed candidate genes were identified: the down-regulated genes AcCRD1 and AcPOR1 involved in chlorophyll biosynthesis, and the up-regulated genes AcSGR1 and AcSGR2 driving chlorophyll degradation. Prochlorophyllide and chlorophyllide, the metabolites produced by AcCRD1 and AcPOR1, progressively reduced in 'JS', but not in 'XX', indicating that chlorophyll biosynthesis was less active in yellow-fleshed fruit. AcSGR1 and AcSGR2 were verified to be involved in chlorophyll degradation, using both transient expression in tobacco and stable overexpression in kiwifruit. Furthermore, a homeobox-leucine zipper (HD-Zip II), AcHZP45, showed significantly increased expression during 'JS' fruit ripening, which led to both repressed expression of AcCRD1 and AcPOR1 and activated expression of AcSGR1 and AcSGR2. Collectively, the present study indicated that different dynamics of chlorophyll biosynthesis and degradation coordinate the changes in chlorophyll content in kiwifruit flesh, which are orchestrated by the key transcription factor AcHZP45.

Comparative biochemical and transcriptomic analysis reveals the phosphate-starving tolerance of two ramie varieties

Ramie (Boehmeria nivea L.) is a highly valued fiber crop. Its yield is often limited by lack of available phosphate (Pi) in the soil, but the underlying molecular mechanisms of ramie's response to Pi deficiency remain largely unknown. To investigate how ramie adapts to low Pi stress, we selected a low Pi-tolerant variety (H-5) and a low Pi-sensitive variety (XYL), and conducted a biochemical and transcriptomic analysis on roots and leaves of both varieties. After subjecting the plants to Pi-deficient and Pi-sufficient conditions for 15 days, we found that H-5 exhibited higher dry weight, longer root systems, and higher levels of Pi, galactolipids, and organic acids when subjected to Pi deprivation, compared to XYL. Transcriptomic analysis further revealed that Pi-responsive genes involved in lipid metabolism, Pi transport, organic acid synthesis, and acid phosphatase activities were more induced in the tolerant variety H-5. Furthermore, weighted gene co-expression network analysis (WGCNA) identified five hub genes, including phosphate transporter, SPX domain-containing protein and sulfoquinovosyl transferase, which played key roles in low Pi tolerance in ramie. The present study will broaden our comprehension of the differences and molecular mechanisms of different ramie cultivars in response to Pi starvation, and lay a foundation for future agronomic improvements in ramie and other fiber crops.

Comparative biochemical kinase activity analysis identifies rivoceranib as a highly selective VEGFR2 inhibitor

Vascular endothelial growth factor receptor 2 (VEGFR2), a key regulator of tumor angiogenesis, is highly expressed across numerous tumor types and has been an attractive target for anti-cancer therapy. However, clinical application of available VEGFR2 inhibitors has been challenged by limited efficacy and a wide range of side effects, potentially due to inadequate selectivity for VEGFR2. Thus, development of potent VEGFR2 inhibitors with improved selectivity is needed. Rivoceranib is an orally administered tyrosine kinase inhibitor that potently and selectively targets VEGFR2. A comparative understanding of the potency and selectivity of rivoceranib and approved inhibitors of VEGFR2 is valuable to inform rationale for therapy selection in the clinic. Here, we performed biochemical analyses of the kinase activity of VEGFR2 and of a panel of 270 kinases to compare rivoceranib to 10 FDA-approved kinase inhibitors (“reference inhibitors”) with known activity against VEGFR2. Rivoceranib demonstrated potency within the range of the reference inhibitors, with a VEGFR2 kinase inhibition IC50 value of 16 nM. However, analysis of residual kinase activity of the panel of 270 kinases showed that rivoceranib displayed greater selectivity for VEGFR2 compared with the reference inhibitors. Differences in selectivity among compounds within the observed range of potency of VEGFR2 kinase inhibition are clinically relevant, as toxicities associated with available VEGFR2 inhibitors are thought to be partly due to their effects against kinases other than VEGFR2. Together, this comparative biochemical analysis highlights the potential for rivoceranib to address clinical limitations associated with off-target effects of currently available VEGFR2 inhibitors.

Abstract 4014: Comparative biochemical kinase activity analysis identifies rivoceranib as the most selective VEGFR-2 inhibitor compared with other TKIs with known activity against VEGFR-2

Abstract Introduction: Vascular endothelial growth factor receptor 2 (VEGFR-2) is a key regulator of tumor angiogenesis that is highly expressed in several tumor types and is a known target for anti-cancer therapy. Yet the clinical use of VEGFR-2 inhibitors has been challenged by limited efficacy and various side effects, potentially due to the low selectivity of these TKIs for VEGFR-2. Thus, potent VEGFR-2 inhibitors with improved selectivity are needed. Rivoceranib is an oral tyrosine kinase inhibitor (TKI) that potently and selectively inhibits VEGFR-2. A comparison of the potency and selectivity of VEGFR-2 inhibitors can provide a rationale for selecting a specific TKI for anticancer therapy in the clinic. Methods: Binding of rivoceranib to VEGFR-2 was determined on a Biacore T200. The affinity constant (KD) was derived from the association and dissociation rate constants. Inhibitory potency of rivoceranib and 10 FDA-approved reference inhibitors on kinase enzyme activity was determined using mobility shift assays (MSA) or immobilized metal ion affinity particle (IMAP) assays. The half-maximum inhibitory activity (IC50) of the 11 inhibitors on VEGFR-2 was determined in 10-point dose-response curves. The selectivity of the inhibitors was determined on 270 wild-type kinases at a fixed concentration of each inhibitor. Rivoceranib was tested at 10- and 100-times IC50 (160 nmol/L and 1.6 µmol/L). Reference inhibitors were tested at 1 µmol/L (35 to 1056-times IC50). Results: Rivoceranib had a KD of 3 nmol/L on VEGFR-2. In enzyme activity assays, rivoceranib had intermediate potency compared with the 10 reference inhibitors, with a VEGFR-2 kinase inhibition IC50 value of 16 nmol/L. Analysis of the residual activity of the panel of 270 kinases in the presence of rivoceranib or the reference inhibitors showed wide variation in selectivity for VEGFR-2, with rivoceranib identified as the most selective inhibitor (activity of 16 additional kinases inhibited by &amp;gt;50% at 1.6 µmol/L). Tivozanib, the most potent VEGFR-2 inhibitor, displayed greater than 50% inhibitory activity against more than 70 additional kinases. Sunitinib was identified as the least selective inhibitor included in this study, inhibiting 125 additional kinases by &amp;gt;50%. Conclusion: Variations in selectivity among TKIs with similar anti-VEGFR-2 potency can help explain differences in their clinical toxicity profiles, which may be partially due to variant inhibitory effects against TKIs other than VEGFR-2. This comparative biochemical analysis highlights the potential for rivoceranib to address clinical limitations associated with the poor selectivity of currently available VEGFR-2 inhibitors. Rivoceranib is under ongoing investigation as monotherapy and in combination with chemotherapy in various tumor types. Citation Format: Seong Jang, Bill Strickland, Lynda Finis, Jeffrey J. Koojiman, Janneke J. Melis, Guido J. Zaman, Jan V. Tornout. Comparative biochemical kinase activity analysis identifies rivoceranib as the most selective VEGFR-2 inhibitor compared with other TKIs with known activity against VEGFR-2. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4014.

Comparative Biochemical and Transcriptome Analyses Reveal Potential Candidate Genes Related to Pericarp Browning in Red Rice

Proanthocyanidins (PAs) are major phytochemicals in rice, and accumulate abundantly in red pericarp (RP) rice. Consumers and rice breeders are increasingly showing interest in PAs because of their beneficial health effects; however, PA biosynthesis in rice is not well-understood. Therefore, to gain insights into this process, we performed comparative transcriptome analysis of grains harvested at 14 days after flowering (DAF; i.e., the stage at which active PA biosynthesis occurs without pericarp color change) and 28 DAF (the stage of late seed development with pericarp color change) from RP and white pericarp rice. In RP rice at 14 DAF, the expression levels of six structural genes (OsCHS, OsF3H, OsF3′H, OsDFR, OsANS, and OsLAR), one modification gene (OsUGT), and one transport gene (OsGSTU34) were significantly upregulated, along with the activation of Rc, the key regulator of PA accumulation in the pericarp. Functional enrichment analysis of 56 differentially expressed genes specifically upregulated in RP rice at 28 DAF revealed the presence of three laccase genes known to cause the browning reaction through oxidation. These results expand our understanding of PA biosynthesis in rice, and provide a genetic basis that will lead to further studies on the genes and underlying molecular mechanisms associated with this process.

Comparative studies of effect of compound feeds on fish-breeding and biochemical properties of juvenile chum salmon

Effectiveness of starter fodders is examined for the low-temperature rearing pools in the salmon fish hatchery Savushkina (Severo-Kurilsk). In May 2022, the experimental compound feed developed in TINRO was compared with the control fodder Aller Aqua (made in Denmark) when feeding of chum salmon fry. Both fodders complied with the veterinary and sanitary requirements of the Euro-Asian Economic Union in terms of quality and safety. The following fish breeding indices were determined for the experimental/control fodders after 47 days of testing: feeding ratio 0.670/0.467; mean daily gain 1.255/1.547 %; mean final weight 0.630/0.735 g; absolute weight growth 0.287/0.392 g; mortality (survival) 0.50 (99.50)/0.48 (99.52) %. Increasing of the water and minerals content and noticeable decreasing of the protein and fat content were detected in tissues of the control samples of fry by comparative biochemical analysis before and after the testing, whereas the content of protein and minerals had increased in tissues of the experimental samples. Thus, suitability and effectiveness of the experimental starter fodder for rearing of juvenile chum salmon in the low-temperature environment is proven by combination of several indices. To intensify the weight gain of juveniles, the experiments will be continued to adjust the composition of mixed feed for juvenile chum salmon in the cold-water conditions, with a principal restriction of the growth intensifiers (hormones) use. The starter compound feed developed in TINRO is recommended for using in rearing of chum salmon fry.

Structural and Functional Insights into the Roles of Potential Metal-Binding Sites in Apostichopus japonicus Ferritin.

Ferritin is widely acknowledged as a conservative iron storage protein found in almost all living kingdoms. Apostichopus japonicus (Selenka) is among the oldest echinoderm fauna and has unique regenerative potential, but the catalytic mechanism of iron oxidation in A. japonicus ferritin (AjFER) remains elusive. We previously identified several potential metal-binding sites at the ferroxidase center, the three- and four-fold channels in AjFER. Herein, we prepared AjFER, AjFER-E25A/E60A/E105A, AjFER-D129A/E132A, and AjFER-E168A mutants, investigated their structures, and functionally characterized these ferritins with respect to Fe2+ uptake using X-ray techniques together with biochemical analytical methods. A crystallographic model of the AjFER-D129A/E132A mutant, which was solved to a resolution of 1.98 Å, suggested that the substitutions had a significant influence on the quaternary structure of the three-fold channel compared to that of AjFER. The structures of these ferritins in solution were determined based on the molecular envelopes of AjFER and its variants by small-angle X-ray scattering, and the structures were almost consistent with the characteristics of well-folded and globular-shaped proteins. Comparative biochemical analyses indicated that site-directed mutagenesis of metal-binding sites in AjFER presented relatively low rates of iron oxidation and thermostability, as well as weak iron-binding affinity, suggesting that these potential metal-binding sites play critical roles in the catalytic activity of ferritin. These findings provide profound insight into the structure-function relationships related to marine invertebrate ferritins.

Comparative Biochemical Profiling of Aluminum Chloride and Sodium Azide Induced Neuroinflammation and Cardiometabolic Disturbance

Comorbidities in human beings signify the numerous risk factors that increase the incidences of neuro- and cardio-metabolic disorders. Experimental models depicting comorbidities are important to explore the molecular pathophysiology that can help suggest appropriate treatment strategies. Tissue-accumulating potential and pathological effects of aluminium chloride (AlCl3)and sodium azide (NaN3) are well recognized. Hence, in the current work, we have for the first time aimed to investigate the unexplored potential of graded dose effects of AlCl3 and NaN3 in inducing early inflammation and cardiometabolic toxicity via comparative biochemical analysis of AlCl3 and/or NaN3. Rats were allocated into seven groups (n = 6). Group 1 was normal control. Remaining groups were given graded doses of AlCl3 and/or NaN3, as LD-AlCl3 (AlCl3 40 mg), MD-AlCl3 (AlCl3 45 mg), and HD-AlCl3 (AlCl3 50 mg) representing low dose, medium dose, and high dose of AlCl3, respectively, and the remaining as LD-NaN3 (NaN3 13 mg), MD-NaN3 (NaN3 15 mg), and HD-NaN3 (NaN3 17 mg) representing low dose, medium dose, and high dose of NaN3, respectively. Serum levels of glucose, insulin, lipid profile, inflammatory mediators like IL-6 and oxidative stress marker, and malondialdehyde (MDA) were analyzed. Likewise, subacute toxicity parameters were analyzed. Immunohistochemistry (IHC) and histopathology (H&E/Masson's trichrome staining) of brain, heart, and pancreatic tissues were done. ECG pattern of all groups was observed. HD-AlCl3 was associated with elevated levels of inflammatory biomarkers, MDA, and glycemic and lipid profiles, whereas it decreased the insulin levels. HD-NaN3 also showed the similar effects of aggravated inflammatory biomarkers, impaired glycemic and lipid profiles, but depicted the maximum mortality rate as compared to HD-AlCl3. IHC showed prominent amyloid plaques and neurofibrillary tangle formation with MD-AlCl3 and HD-AlCl3 as compared to NaN3-treated groups. Likewise, in brain tissues, vacuolation of white matter, vascular congestion, and hemorrhage were seen in HD-AlCl3 treated group, while HD-NaN3 induced death in animals. AlCl3 exposure resulted in an inverted QRS complex, while exposure to NaN3 showed ST depression but with increased mortality. AlCl3 has better controlled results as compared to NaN3 for induction of comorbid experimental animal model depicting early neuroinflammation and cardiometabolic disruption. These determined efforts facilitate the researchers for the development of clinically effective treatment strategies using such experimental models.

Structural and functional determinants of the archaeal 8-oxoguanine-DNA glycosylase AGOG for DNA damage recognition and processing.

8-Oxoguanine (GO) is a major purine oxidation product in DNA. Because of its highly mutagenic properties, GO absolutely must be eliminated from DNA. To do this, aerobic and anaerobic organisms from the three kingdoms of life have evolved repair mechanisms to prevent its deleterious effect on genetic integrity. The major way to remove GO is the base excision repair pathway, usually initiated by a GO-DNA glycosylase. First identified in bacteria (Fpg) and eukaryotes (OGG1), GO-DNA glycosylases were more recently identified in archaea (OGG2 and AGOG). AGOG is the less documented enzyme and its mode of damage recognition and removing remains to be clarified at the molecular and atomic levels. This study presents a complete structural characterisation of apo AGOGs from Pyrococcus abyssi (Pab) and Thermococcus gammatolerans (Tga) and the first structure of Pab-AGOG bound to lesion-containing single- or double-stranded DNA. By combining X-ray structure analysis, site directed mutagenesis and biochemistry experiments, we identified key amino acid residues of AGOGs responsible for the specific recognition of the lesion and the base opposite the lesion and for catalysis. Moreover, a unique binding mode of GO, involving double base flipping, never observed for any other DNA glycosylases, is revealed. In addition to unravelling the properties of AGOGs, our study, through comparative biochemical and structural analysis, offers new insights into the evolutionary plasticity of DNA glycosylases across all three kingdoms of life.

Comparative Biochemical and Transcriptomic Analyses Provide New Insights into Phytoplasma Infection Responses in Cucumber.

Flat stem and witches’ broom phytoplasma-like symptoms in the cucumber inbred line C17 were observed in a greenhouse at Yangzhou University, China for three consecutive planting seasons; these symptoms resulted in a decreased yield. To better understand the cause of these symptoms, 16S rRNA PCR, plant hormones, mineral elements, and RNA-seq profiling were performed using symptomatic and normal stem samples. The results showed that the causal agent was classified as the Candidatus phytoplasma asteris strain, a plant pathogenic prokaryote that could not be cultured in vitro. Measurement of plant hormones showed that auxin, salicylic acid, and jasmonic acid contents were significantly increased, whereas that of ethylene’s immediate biosynthetic precursor, 1-aminocyclopropane-1-carboxylic acid, was decreased in the phytoplasma-infected stems compared with the healthy stems. Furthermore, measurement of mineral element composition showed that magnesium, calcium, sodium, iron, and zinc concentrations significantly changed in the phytoplasma-infected cucumber stems compared with the uninfected stems. Comparative RNA-seq identified 253 differentially expressed genes, including 179 upregulated and 74 downregulated genes. Further analyses suggested that genes related to phenylpropanoid biosynthesis, phenylalanine metabolism, and plant hormone signal transduction contributed to phytoplasma infection. Taken together, this study presents the first in-depth assessment of disease symptoms and biochemical content of cucumber stems known to be infected with phytoplasma.

Biochemical Analysis of Urine Samples from Diabetic and Hypertensive Patients without Renal Dysfunction Using Spectrophotometry and Raman Spectroscopy Techniques Aiming Classification and Diagnosis.

The purpose of this study was to perform a comparative biochemical analysis between conventional spectrophotometry and Raman spectroscopy, techniques used for diagnoses, on the urine of healthy (CT) and diabetic and hypertensive patients (DM&HBP). Urine from 40 subjects (20 in the CT group and 20 in the DM&HBP group) was examined in a dispersive Raman spectrometer (an 830 nm excitation and a 350 mW power). The mean Raman spectra between both groups showed a significant difference in peaks of glucose; exploratory analysis by principal component analysis (PCA) identified spectral differences between the groups, with higher peaks of glucose and proteins in the DM&HBP group. A partial least squares (PLS) regression model estimated by the Raman data indicated the concentrations of urea, creatinine, glucose, phosphate, and total protein; creatinine and glucose were the biomarkers that presented the best correlation coefficient (r) between the two techniques analyzed (r = 0.68 and r = 0.98, respectively), both with eight latent variables (LVs) and a root mean square error of cross-validation (RMSecv) of 3.6 and 5.1 mmol/L (41 and 92 mg/dL), respectively. Discriminant analysis (PLS-DA) using the entire Raman spectra was able to differentiate the samples of the groups in the study, with a higher accuracy (81.5%) compared to the linear discriminant analysis (LDA) models using the concentration values of the spectrometric analysis (60.0%) and the concentrations predicted by the PLS regression (69.8%). Results indicated that spectral models based on PLS applied to Raman spectra may be used to distinguish subjects with diabetes and blood hypertension from healthy ones in urinalysis aimed at population screening.

Heterologous expression and biochemical comparison of two homologous SoxX proteins of endosymbiotic Candidatus Vesicomyosocius okutanii and free-living Hydrogenovibrio crunogenus from deep-sea environments

Candidatus Vesicomyosocius okutanii is a currently uncultured endosymbiotic bacterium of Phreagena okutanii, a clam that inhabits deep-sea vent environments. The genome of Ca. V. okutanii encodes a sulfur-oxidizing (Sox) enzyme complex, presumably generating biological energy for the host from inorganic sulfur compounds. Here, Ca. V. okutanii SoxX (VoSoxX), a mono-heme cytochrome c component of the Sox complex, was shown to be phylogenetically related to its homologous counterpart (HcSoxX) from a free-living deep-sea bacterium, Hydrogenovibrio crunogenus. Both proteins were heterologously expressed in Escherichia coli co-expressing cytochrome c maturation genes for comparative biochemical analysis. The VoSoxX recombinant had significantly lower thermal stability than HcSoxX, reflecting the difference in growth conditions of the source bacteria. The endosymbiont inhabits a mild intracellular environment, whereas the free-living bacterium dwells in a harsh environment. This study represents the first successful case of heterologous expression of genes from Ca. V. okutanii, allowing further biochemical studies of the molecular mechanism of sulfur oxidation in deep-sea environments.