Ferroxidase Activity Research Articles

Structural and functional relationship of mammalian and nematode ferritins.

Ferritin is a unique buffering protein in iron metabolism. By storing or releasing iron in a tightly controlled manner, it prevents the negative effects of free ferrous ions on biomolecules in all domains of life - from bacteria to mammals. This review focuses on the structural features and activity of the ferritin protein family with an emphasis on nematode ferritins and the similarities in their biological roles with mammalian ferritins. The conservative characteristic of the ferritin family across the species originates from the ferroxidase activity against redox-active iron. The antioxidative function of these proteins translates into their involvement in a wide range of important biological processes, e.g., aging, fat metabolism, immunity, anticancer activity, and antipathogenic activity. Moreover, disturbances in ferritin expression lead to severe iron-associated diseases. Research on the Caenorhabditis elegans model organism may allow us to better understand the wide spectrum of mechanisms involving ferritin activity.

Expression of soluble, active, fluorescently tagged hephaestin in COS and CHO cell lines.

Hephaestin (Hp) is a trans-membrane protein, which plays a critical role in intestinal iron absorption. Hp was originally identified as the gene responsible for the phenotype of sex-linked anaemia in the sla mouse. The mutation in the sla protein causes accumulation of dietary iron in duodenal cells, causing severe microcytic hypochromic anaemia. Although mucosal uptake of dietary iron is normal, export from the duodenum is inhibited. Hp is homologous to ceruloplasmin (Cp), a member of the family of multi copper ferroxidases (MCFs) and possesses ferroxidase activity that facilitates iron release from the duodenum and load onto the serum iron transport protein transferrin. In the present study, attempts were made to produce biologically active recombinant mouse hephaestin as a secretory form tagged with green fluorescent protein (GFP), Hpsec-GFP. Plasmid expressing Hpsec-GFP was constructed and transfected into COS and CHO cells. The GFP aided the monitoring expression in real time to select the best conditions to maximise expression and provided a tag for purifying and analysing Hpsec-GFP. The protein had detectable oxidase activity as shown by in-gel and solution-based assays. The methods described here can provide the basis for further work to probe the interaction of hephaestin with other proteins using complementary fluorescent tags on target proteins that would facilitate the fluorescence resonance energy transfer measurements, for example with transferrin or colocalisation studies, and help to discover more about hephaestin works at the molecular level.

Oxidant and antioxidant balance in children with bacteremia.

There is a crucial balance between oxidant and antioxidant defense mechanisms. We aimed to evaluate the role of the balance of these systems in children with bloodstream infection. We analyzed prospectively oxidant and antioxidant stress parameters from serum samples of children with BSI besides demographic and clinical data of children. Serum levels of the total antioxidant status (TAS), total oxidant status (TOS), albumin, plasma thiol, disulphide, catalase (CAT), myeloperoxidase (MPO), ischemia-modified albumin (IMA) levels, ferroxidase and arylesterase (ARES) activity were evaluated in both patients and healthy controls. A total of 113 children were evaluated, 50 of them had bacteremia and the remaining 63 were healthy subjects. The median TOS values were 18.5 μmol H2O2/L and 13.1 μmol H2O2/L in patient and control groups, respectively with a statistically significant difference between groups. The mean serum IMA levels were 0.8 ± 0.1 absorbance unit (ABSU) in patients and 0.5 ± 0.09 ABSU in control, the difference between groups was statistically significant. The native thiol, total thiol levels and the disulphide levels were significantly lower in the patient group as compared with the control group. The myeloperoxidase level was 136 U/L in patients and 107 in controls with a statistically significant difference between groups. TOS, IMA, MPO, and particularly plasma thiols seem good candidates for accurate diagnosis of bacteremia in children.

Oxidative stress parameters in patients with ascending aortic dilatation

Background/aimThis study aimed to determine plasma thiol, disulphide, and serum ischemia-modified albumin (IMA) levels and ferroxidase activity in patients with ascending aorta dilatation (AAD) in comparison to those without AAD and to evaluate the predictive value of these oxidative stress parameters for AAD.Materials and methods This study was designed as a cross-sectional study of 184 patients who applied to our cardiology clinic. Our study population consisted of patients with AAD (n = 85) and without AAD (n = 99). A spectrophotometric method was used to determine plasma thiol, disulphide, and serum IMA levels and ferroxidase activity.Results The native thiol and the total thiol levels were significantly higher in the control group than the AAD group (P < 0.001), whereas the disulphide and IMA levels and the ferroxidase activity were similar between the groups. The native thiol and the total thiol levels were inversely and significantly correlated with ascending aortic diameter (r = –0.38, P < 0.001; r = –0.39, P < 0.001; respectively). The left ventricle mass and the total thiol levels were independent predictors of ascending aortic diameter (β= 0.223, P = 0.02; β= –0.340, P < 0.001; respectively).Conclusion Among oxidative stress parameters including thiols, disulphide, IMA, and ferroxidase activity, only the lower total thiol levels appear to confer a high risk for AAD development. Along with the proven diagnostic imaging methods, thiol levels may be helpful to diagnose and stratify patients with AAD.

Effects of Dietary Glucose and Fructose on Copper, Iron, and Zinc Metabolism Parameters in Humans.

Alterations of transition metal levels have been associated with obesity, hepatic steatosis, and metabolic syndrome in humans. Studies in animals indicate an association between dietary sugars and copper metabolism. Our group has conducted a study in which young adults consumed beverages sweetened with glucose, fructose, high fructose corn syrup (HFCS), or aspartame for two weeks and has reported that consumption of both fructose- and HFCS-sweetened beverages increased cardiovascular disease risk factors. Baseline and intervention serum samples from 107 participants of this study were measured for copper metabolism (copper, ceruloplasmin ferroxidase activity, ceruloplasmin protein), zinc levels, and iron metabolism (iron, ferritin, and transferrin) parameters. Fructose and/or glucose consumption were associated with decreased ceruloplasmin ferroxidase activity and serum copper and zinc concentrations. Ceruloplasmin protein levels did not change in response to intervention. The changes in copper concentrations were correlated with zinc, but not with iron. The decreases in copper, ceruloplasmin ferroxidase activity, ferritin, and transferrin were inversely associated with the increases in metabolic risk factors associated with sugar consumption, specifically, apolipoprotein CIII, triglycerides, or post-meal glucose, insulin, and lactate responses. These findings are the first evidence that consumption of sugar-sweetened beverages can alter clinical parameters of transition metal metabolism in healthy subjects.

The ferroxidase LPR5 functions in the maintenance of phosphate homeostasis and is required for normal growth and development of rice.

Members of the Low Phosphate Root (LPR) family have been identified in rice (Oryza sativa) and expression analyses have been conducted. Here, we investigated the functions of one of the five members in rice, LPR5. qRT-PCR and promoter-GUS reporter analyses indicated that under Pi-sufficient conditions OsLPR5 was highly expressed in the roots, and specific expression occurred in the leaf collars and nodes, and its expression was increased under Pi-deficient conditions. In vitro analysis of the purified OsLPR5 protein showed that it exhibited ferroxidase activity. Overexpression of OsLPR5 triggered higher ferroxidase activity, and elevated concentrations of Fe(III) in the xylem sap and of total Fe in the roots and shoots. Transient expression of OsLPR5 in Nicotiana benthamiana provided evidence of its subcellular localization to the cell wall and endoplasmic reticulum. Knockout mutation in OsLPR5 by means of CRISPR-Cas9 resulted in adverse effects on Pi translocation, on the relative expression of Cis-NATOsPHO1;2, and on several morphological traits, including root development and yield potential. Our results indicate that ferroxidase-dependent OsLPR5 has both a broad-spectrum influence on growth and development in rice as well as affecting a subset of physiological and molecular traits that govern Pi homeostasis.

Two-photon imaging of oxidative stress in living erythrocytes as a measure for human aging.

According to the "oxidative stress theory" of aging, this process is accompanied by a progressive and irreversible accumulation of oxidative damage caused by reactive oxygen species (ROS). This, in turn, has a deleterious impact on molecular mechanisms in aging thereby altering the physiological function of the organism, increasing the risk of different aging-related diseases, as well as impacting the life span. The aim of the current study was to investigate oxidative stress in living red blood cells (RBCs) in human aging as an oxidative stress-related pathological condition. Two-photon laser scanning and light microscopy techniques were applied to analyze the oxidative stress in RBCs and the cell viability. Spectrophotometric analyzes were performed to determine the percentage of RBC hemolysis, activities of superoxide dismutase and catalase in RBCs, as well as the ferroxidase activities of ceruloplasmin in blood plasma samples. The studies included three human aging groups, young, middle-aged, and elderly. According to the results, the two-photon fluorescence of carboxy-DCFDA, indicating the intensity of oxidative stress, significantly increase in RBCs by the increase of age (P < 0.05), and these intensities are in statistically significant positive correlation with age (P < 0.001) and a strong negative correlation (P < 0.05) with the activity of catalase in RBCs and ferroxidase activity of ceruloplasmin in plasma. In conclusion, two-photon fluorescent imaging of oxidative stress in human living RBCs is a valuable and accurate method for the determination of aging processes in humans and can be suggested as a novel indicator for human aging processes in individual aging.

Deletion of H-ferritin in macrophages alleviates obesity and diabetes induced by high-fat diet in mice.

Iron accumulation affects obesity and diabetes, both of which are ameliorated by iron reduction. Ferritin, an iron-storage protein, plays a crucial role in iron metabolism. H-ferritin exerts its cytoprotective action by reducing toxicity via its ferroxidase activity. We investigated the role of macrophage H-ferritin in obesity and diabetes. Conditional macrophage-specific H-ferritin (Fth, also known as Fth1) knockout (LysM-Cre Fth KO) mice were used and divided into four groups: wild-type (WT) and LysM-Cre Fth KO mice with normal diet (ND), and WT and LysM-Cre Fth KO mice with high-fat diet (HFD). These mice were analysed for characteristics of obesity and diabetes, tissue iron content, inflammation, oxidative stress, insulin sensitivity and metabolic measurements. RAW264.7 macrophage cells were used for in vitro experiments. Iron concentration reduced, and mRNA expression of ferroportin increased, in macrophages from LysM-Cre Fth KO mice. HFD-induced obesity was lower in LysM-Cre Fth KO mice than in WT mice at 12weeks (body weight: KO 34.6 ± 5.6g vs WT 40.1 ± 5.2g). mRNA expression of inflammatory cytokines and infiltrated macrophages and oxidative stress increased in the adipose tissue of HFD-fed WT mice, but was not elevated in HFD-fed LysM-Cre Fth KO mice. However, WT mice fed an HFD had elevated iron concentration in adipose tissue and spleen, which was not observed in LysM-Cre Fth KO mice fed an HFD (adipose tissue [μmol Fe/g protein]: KO 1496 ± 479 vs WT 2316 ± 866; spleen [μmol Fe/g protein]: KO 218 ± 54 vs WT 334 ± 83). Moreover, HFD administration impaired both glucose tolerance and insulin sensitivity in WT mice, which was ameliorated in LysM-Cre Fth KO mice. In addition, energy expenditure, mRNA expression of thermogenic genes, and body temperature were higher in KO mice with HFD than WT mice with HFD. In vitro experiments showed that iron content was reduced, and lipopolysaccharide-induced Tnf-α (also known as Tnf) mRNA upregulation was inhibited in a macrophage cell line transfected with Fth siRNA. Deletion of macrophage H-ferritin suppresses the inflammatory response by reducing intracellular iron levels, resulting in the prevention of HFD-induced obesity and diabetes. The findings from this study highlight macrophage iron levels as a potential therapeutic target for obesity and diabetes.

Effect of Narrowband-UVB Phototherapy Treatment on Total Ferroxidase Activity and Ceruloplasmin Oxidase Activity in Sera of Iraqi Vitiligo Patients

Vitiligo is an acquired pigmentary disorder in which white depigmented macules and patches of skin appear on different parts of the body, affecting all ages and both sexes equally. Focal, segmen-tal, generalized and universal are considered the most characteristic patterns of vitiligo. One of the most effective treatment of this disorder is using light therapy, and in particular ultraviolet (UV) light. Among them narrowband ultraviolet B phototherapy (NB-UVB) is the most widely used. Oxidative stress is considered to be one of the possible pathogenic events in melanocyte loss. Imbalance in the oxidant/antioxidant system have been demonstrated in blood of vitiligo patients. To demonstrate the safety of using NB-UVB phototherapy treatment in vitiligo Iraqi patients. Patients with vitiligo group were treated with NB-UVB regimen as three times weekly on non- consecutive days for a total of 20 sessions (about 2 months). Total ferroxidase (activity and specific activity), ceruloplasmin concentration, ceruloplasmin oxidase (activity and specific activity) and serum copper were determined in all participants before and after NBUVB phototherapy. The above biochemical parameters were measured in serum samples of 30 vitiligo patients before and after treatment with the narrow band UVB phototherapy. NB-UVB treatment was found to be associated with a significant reduction in copper ion concentration, but no significant differences in total ferroxidase (activity and specific activity), ceruloplasmin oxidase (activity and specific activity), and ceruloplasmin concentration. The present results provide evidence which support using narrow band UVB phototherapy as a safe method for vitiligo treatment.

Rapid Decay of the Native Intermediate in the Metallooxidase Fet3p Enables Controlled FeII Oxidation for Efficient Metabolism.

The multicopper oxidases (MCOs) couple four 1e- oxidations of substrate to the 4e- reduction of O2 to H2O. These divide into two groups: those that oxidize organic substrates with high turnover frequencies (TOFs) up to 560 s-1 and those that oxidize metal ions with low TOFs, ∼1 s-1 or less. The catalytic mechanism of the organic oxidases has been elucidated, and the high TOF is achieved through rapid intramolecular electron transfer (IET) to the native intermediate (NI), which only slowly decays to the resting form. Here, we uncover the factors that govern the low TOF in Fet3p, a prototypical metallooxidase, in the context of the MCO mechanism. We determine that the NI decays rapidly under optimal turnover conditions, and the mechanism thereby becomes rate-limited by slow IET to the resting enzyme. Development of a catalytic model leads to the important conclusions that proton delivery to the NI controls the mechanism and enables the slow turnover in Fet3p that is functionally significant in Fe metabolism enabling efficient ferroxidase activity while avoiding ROS generation.

Dps protein is related to resistance of Mycobacterium abscessus subsp. massiliense against stressful conditions.

Mycobacterium abscessus subsp. massiliense (Mycma) belongs to the Mycobacterium abscessus complex and is a rapidly growing non-tuberculous mycobacterium. The chronic pulmonary, skin, and soft tissue infections that it causes may be difficult to treat due to its intrinsic resistance to the commonly used antimicrobial drugs, making it a serious world public health problem. Iron is an essential nutrient for the growth of microorganisms; nonetheless, it can be toxic when in excess. Thus, bacteria require an iron homeostasis mechanism to succeed in different environments. DNA-binding proteins from starved cells (Dps) are miniferritins with the property to act as additional iron storage proteins but also can bind to DNA, protecting it against hydroxyl radical. Annotation of the Mycma genome revealed the gene mycma_03135 with 79% sequential identity when compared to MSMEG_3242 gene from M. smegmatis mc2 155, which codifies for a known Dps. Recombinant Dps from M. abscessus (rMaDps) was produced in Escherichia coli, purified in soluble form and shown to form high mass oligomers in solution with ferroxidase activity, DNA binding, and protection against damage. The expression of the mycma_03135 gene was induced during Mycma growth in the presence of hydrogen peroxide (H2O2). Additionally, the expression of rMaDps by E. coli conferred greater resistance to H2O2. Thus, this study is the first to identify and characterize a Dps from M. abscessus. KEY POINTS: Mycobacterium abscessus subsp. massiliense express a miniferritin protein (Dps). Mycma Dps binds to DNA and protects against oxidative stress.

Analysis of concentration and activity of proteins involved in iron metabolism in rats with streptozotocin-induced hyperglycemia

Objective. We aimed to analyze the alterations of concentration and activity of iron metabolism proteins in samples obtained from rats with hyperglycemia induced by streptozotocin (STZ).&#x0D; Materials and methods. Concentration and activity of ceruloplasmin (Cp) and transferrin (Tf), concentration of glucose, fructosamine, hemoglobin, ferritin (Ft), and iron were measured in blood samples obtained from rats after injection of STZ or saline (control). To develop in-house ELISA, highly purified preparations of Cp, Tf, and Ft, as well as specific antibodies against these proteins, were obtained.&#x0D; Results. The hyperglycemia in rats after STZ injection was confirmed by increase in glucose and fructosamine serum concentrations. Increase in Cp concentration and decrease in specific ferroxidase activity of Cp, concentration of Tf and Ft were observed in hyperglycemic rats. However, the absence of changes in iron concentration and total iron binding capacity of serum is indicative of compensatory response.&#x0D; Conclusion. STZ-induced hyperglycemia in rats was characterized by alteration of activity and concentration of iron metabolism proteins, however, this alteration was negated by homeostatic response. The alterations of Cp and Tf concentrations observed in this study are similar to those in acute phase of inflammation.

Genetic and Clinical Heterogeneity in Thirteen New Cases with Aceruloplasminemia. Atypical Anemia as a Clue for an Early Diagnosis.

Aceruloplasminemia is a rare autosomal recessive genetic disease characterized by mild microcytic anemia, diabetes, retinopathy, liver disease, and progressive neurological symptoms due to iron accumulation in pancreas, retina, liver, and brain. The disease is caused by mutations in the Ceruloplasmin (CP) gene that produce a strong reduction or absence of ceruloplasmin ferroxidase activity, leading to an impairment of iron metabolism. Most patients described so far are from Japan. Prompt diagnosis and therapy are crucial to prevent neurological complications since, once established, they are usually irreversible. Here, we describe the largest series of non-Japanese patients with aceruloplasminemia published so far, including 13 individuals from 11 families carrying 13 mutations in the CP gene (7 missense, 3 frameshifts, and 3 splicing mutations), 10 of which are novel. All missense mutations were studied by computational modeling. Clinical manifestations were heterogeneous, but anemia, often but not necessarily microcytic, was frequently the earliest one. This study confirms the clinical and genetic heterogeneity of aceruloplasminemia, a disease expected to be increasingly diagnosed in the Next-Generation Sequencing (NGS) era. Unexplained anemia with low transferrin saturation and high ferritin levels without inflammation should prompt the suspicion of aceruloplasminemia, which can be easily confirmed by low serum ceruloplasmin levels. Collaborative joint efforts are needed to better understand the pathophysiology of this potentially disabling disease.

Structural Insight into Binary Protein Metal-Organic Frameworks with Ferritin Nanocages as Linkers and Nickel Clusters as Nodes.

Metal-organic frameworks (MOFs) hold great promise for numerous applications. However, proteins, carriers of biological functions in living systems, have not yet been fully explored as building blocks for the construction of MOFs. This work presents a strategy for the fabrication of binary MOFs. Considering octahedral ferritin symmetry, four His2 (His-His) motifs were first incorporated into the exterior surface of a ferritin nanocage near each C4 channel, yielding protein linkers with multiple metal-binding sites (bisH-SF). Secondly, by adding nickel ions to bisH-SF solutions triggers the self-assembly of ferritin nanocages into a porous 3D crystalline MOF with designed protein lattice, where two adjacent ferritin molecules along the C4 symmetry axes are bridged by four dinuclear or tetranuclear nickel clusters depending on Ni2+ concentration. This work provides a simple approach for precise control over a binary protein-metal crystalline framework, and the resulting MOFs exhibited inherent ferroxidase activity and peroxidase-like catalytic activity.

‘Silver man’ – A first case report of silver associated steatohepatitis (SASH) and discussion of its pathomechanism

A 47-year-old Chinese man, who worked as a clerk in the electronics industry, presented with gait instability and glove-and-stocking sensation. He had anaemia, deranged liver function, low ceruloplasmin (0.05 g/L; ref 0.18–0.35), high 24-urine copper (2.02 μmol/L; ref <0.5 μmol/L), low iron level (5 umol/L; ref 9–33), high ferritin (4521 pmol/L; ref 52–738) and low transferrin saturation (11%; ref 16–45). MRI liver and pancreas showed homogenous T2 hypointensity, a feature of iron deposition. MRI brain showed artefacts in bilateral globus pallidi that suggested mineralisation. Sequencing of ATP7B gene did not reveal any pathogenic variants. Further clarification of the clinical history revealed that colloidal silver was prescribed by his private practitioner for over a year. His blood silver level was about a thousand times above that of controls. In the liver tissues, there were patchy lymphocytic infiltrates with periportal and focal bridging fibrosis, moderate haemosiderin deposits and presence of silver. Clinical symptoms and biochemical results were improved after silver exposure was stopped. Silver binds with ceruloplasmin and loses its ferroxidase activity, so Fe2+ cannot be oxidised to Fe3+ and cannot be carried by transferrin. This leads to intracellular iron overload in the liver, pancreas and neuronal tissues.

Preparation of platinum nanoparticles using iron(ii) as reductant and photosensitized H2 generation on an iron storage protein scaffold.

The quest for efficient solar-to-fuel conversion has led to the development of numerous homogeneous and heterogeneous systems for photochemical stimulation of 2H+ + 2e− → H2. Many such systems consist of a photosensitizer, an H2-evolving catalyst (HEC), and sacrificial electron donor often with an electron relay between photosensitizer and HEC. Colloidal platinum remains a popular HEC. We report here a novel, simple, and high yield synthesis of Pt nanoparticles (Pt NPs) associated with human heavy chain ferritin (Hfn). The formation of the Pt NPs capitalizes on Hfn's native catalysis of autoxidation of Fe(ii)(aq) (ferroxidase activity). Fe(ii) reduces Pt(ii) to Pt(0) and the rapid ferroxidase reaction produces FeO(OH), which associates with and stabilizes the incipient Pt NPs. This Pt/Fe-Hfn efficiently catalyzes photosensitized H2 production when combined with Eosin Y (EY) as photosensitizer and triethanolamine (TEOA) as sacrificial electron donor. With white light irradiation turnover numbers of 300H2 per Pt, 250H2 per EY were achieved. A quantum yield of 18% for H2 production was obtained with 550 nm irradiation. The fluorescence emission of EY is quenched by TEOA but not by Pt/Fe-Hfn. We propose that the photosensitized H2 production from aqueous TEOA, EY, Pt/Fe-Hfn solution occurs via a reductive quenching pathway in which both the singlet and triplet excited states of EY are reduced by TEOA to the anion radical, EY−˙, which in turn transfers electrons to the Pt/Fe-Hfn HEC. Hfn is known to be a remarkably versatile scaffold for incorporation and stabilization of noble metal and semiconductor nanoparticles. Since both EY and Hfn are amenable to scale-up, we envision further refinements to and applications of this photosensitized H2-generating system.

Trace elements and oxidative stress status in patients with psoriasis

IntroductionPsoriasis is a common, inflammatory skin disease of which etiopathogenesis is still not explained clearly, however in which trace elements and oxidative stress are considered to play a role.AimTo evaluate the serum trace element and oxidative stress levels in patients diagnosed with psoriasis.Material and methodsA total of 87 psoriasis patients and 60 healthy subjects were included in the study. Serum sodium (Na), potassium (K), calcium (Ca), phosphorus (P), magnesium (Mg), iron (Fe), selenium (Se), zinc (Zn), copper (Cu) levels, oxidative stress parameters, ischemia-modified albumin (IMA), catalase (CAT), myeloperoxidase (MPO) and ferroxidase (FOX) activity and an inflammatory marker, C-reactive protein (CRP), were examined in all participants.ResultsIMA, IMA/Albumin (IMA/Alb), CAT, Cu, FOX and CRP levels were found to be significantly higher; Se, Zn and albumin levels were significantly lower in the patient group as compared to the control group. No significant difference was found between groups with regard to Na, K, Ca, P, Mg, Fe and MPO levels.ConclusionsSome trace element levels and oxidant–antioxidant balance were changed in psoriasis patients.

Impact of Phosphate on Iron Mineralization and Mobilization in Nonheme Bacterioferritin B from Mycobacterium tuberculosis.

Ferritins are supramolecular nanocage proteins, which synthesize hydrated ferric oxyhydroxide mineral via protein mediated rapid Fe2+ sequestration and ferroxidase reactions. Ferritin minerals are also associated with a significant amount of phosphate, which contribute toward their structure and reactivity. Like iron, phosphate also regulates the pathogenesis of Mycobacterium tuberculosis (Mtb), which expresses two types of ferritin: heme binding bacterioferritin A (BfrA) and nonheme binding bacterioferritin B (BfrB). Unlike Mtb BfrA, the rapid kinetics and mechanism of ferroxidase activity, and the mineral core formation/dissolution in Mtb BfrB are not well explored. Moreover, the effect of physiological levels of phosphate (0-10 mM) on the synthesis, structure, and reactivity of ferritin mineral core also require investigation in detail. Therefore, the stopped-flow rapid kinetics of ferroxidase activity (ΔA650/Δt) of Mtb BfrB was carried out, which detected a transient intermediate similar to diferric peroxo species as observed in frog and human ferritins. Increasing phosphate concentration increased the initial rate of iron mineralization (ΔA350/Δt) and dissolved O2 consumption (both ∼1.5-2-fold). Phosphate not only decreased the amount of iron loading and size of the BfrB mineral core (both up to 2-fold) but also decreased its crystallinity, resembling the variations observed in the core morphology of different native ferritins. In addition, phosphate inhibited the kinetics of reductive iron mobilization (∼6-8-fold) indicating its influence on the stability of the iron mineral core. Hence, the current work provides the kinetic/mechanistic insight toward the ferroxidase activity in Mtb BfrB, apart from demonstrating the role of phosphate toward the structure/reactivity of its iron mineral.

Insights from molecular dynamics simulation of human ceruloplasmin (ferroxidase enzyme) binding with biogenic monoamines.

Human ceruloplasmin (hCP) is a multi-copper oxidase with ferroxidase and amine oxidase activities. Molecular dynamics simulation (MDS) and docking analysis of biogenic monoamines with ceruloplasmin explain the role of Asp1025, Glu935, Glu272, Glu232 and Glu230 together with the binding site water molecules (referred as conserved water molecules) in the stabilization of neurotransmitter (Serotonin, Norepinephrine and Epinephrine) molecules within the binding cavity of hCP. Conserved water molecules are found at specific positions interacting with the protein structures that have sequence similarity. The ethylamine side chain nitrogen atom (N1) of neurotransmitter molecules interacts with water molecules in the binding cavity formed by Asp1025, Glu935 and Glu232 residues. These residues form an acidic triad mimicking a substrate binding cavity. The hydroxyl groups attached to the catechol ring of epinephrine and norepinephrine have been stabilized by Asp230 and Asp232 residues. Data suggests that the recognition of biogenic amines mediates through the N+(amine) ...Asp1025-His1026-CuCis-His path. The potential recognition path of biogenic monoamines to trinuclear copper cluster supported by active site water molecules (referred as conserved water molecules) is described in this report.

Serum Ceruloplasmin levels in Type 2 Diabetes mellitus – Assessing the cardiovascular risk

Type 2 diabetes mellitus is an endocrinological disease associated with hyperglycemia characterized by both insulin resistance and defective insulin secretion. Cardiovascular diseases are the peak reason for mortality in India, highest rate seen in Tamil Nadu state, from the age of 25 to 69 years. Many studies have revealed the importance of inflammatory pathway playing a pivotal role in the development and progression of diabetic complications. Ceruloplasmin, a copper containing mettalo-enzyme having antioxidant property (e.g ferroxidase activity) is an ideal marker to know the cardiovascular status, glycemic status, dyslipidemia and cancer risk. The purpose of the present study was to evaluate the effects of Body Mass Index (BMI) and Ceruloplasmin levels on the incidence of cardiovascular diseases in Type 2 Diabetes mellitus patients. Fasting Blood glucose, lipid profile which includes serum triglyceride, total cholesterol, LDL, VLDL, HDL and serum ceruloplasmin were investigated using fully automated chemistry analyzer. Diabetic patients were segregated into three groups based on their BMI, which was correlated with inflammatory marker ceruloplasmin to assess the cardiovascular risk. The mean value and standard deviation of ceruloplasminin controls (n=60) and diabetic cases divided into normal weight, over weight and obese (n=20 each) were 38.3±8.45, 36±7.99, 33.2±3.57and34.2±6.24 mg/dl respectively. Ceruloplasmin among the cases and controls were not statistically significant. Serum ceruloplasmin levels and BMI showed no correlation among the group 1 diabetic cases and was not significant (p=0.29), among the group 2diabetic cases it showed weak correlation and was not significant (p=0.85), and it also showed no correlation among the group 3 diabetic cases and was not significant (p=0.65).This study revealed no significant association between serum ceruloplasmin level and BMI in all diabetic group subjects separately.