β-N-methylamino-L-alanine Inhibits Human Catalase Activity: Possible Implications for Neurodegenerative Disease Development

The naturally produced, nonprotein amino acid β- N-methylamino-l-alanine (BMAA) has been proposed as a significant contributor to sporadic neurodegenerative disease development worldwide. However, the existing hypothesized mechanisms of toxicity do not adequately explain the role of BMAA in neurodegenerative disease development. There is evidence for BMAA-induced enzyme inhibition, but the effect of BMAA on human stress response enzymes has received little attention, despite the well-described role of oxidative stress in neurodegenerative disease development. The aim of this study was therefore to investigate the effect of BMAA on human catalase activity and compare it to the known inhibitor 3-amino-1,2,4-triazole. BMAA inhibited human erythrocyte catalase in a cell-free exposure to the same extent as the known inhibitor. Based on enzyme kinetics, the inhibition appears to be noncompetitive, possibly as a result of BMAA binding in the nicotinamide adenine dinucleotide phosphate (NADPH) binding site. BMAA-induced catalase inhibition was also observed in a human cell line culture. We therefore propose that BMAA-induced enzyme inhibition, specifically catalase inhibition, is a mechanism of toxicity that may contribute to the neurotoxicity of BMAA, further supporting the role of BMAA in neurodegenerative disease development.

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Enhanced Production of the Mical Redox Domain for Enzymology and F-actin Disassembly Assays

International Journal of Molecular Sciences ◽

10.3390/ijms22041991 ◽

2021 ◽

Vol 22 (4) ◽

pp. 1991

Author(s):

Jimok Yoon ◽

Heng Wu ◽

Ruei-Jiun Hung ◽

Jonathan R. Terman

Keyword(s):

Nicotinamide Adenine Dinucleotide Phosphate ◽

Actin Dynamics ◽

Specific Substrate ◽

Actin Assembly ◽

Redox Enzymes ◽

Enhanced Production ◽

Reversible Redox ◽

Future Work ◽

Signaling Process

To change their behaviors, cells require actin proteins to assemble together into long polymers/filaments—and so a critical goal is to understand the factors that control this actin filament (F-actin) assembly and stability. We have identified a family of unusual actin regulators, the MICALs, which are flavoprotein monooxygenase/hydroxylase enzymes that associate with flavin adenine dinucleotide (FAD) and use the co-enzyme nicotinamide adenine dinucleotide phosphate (NADPH) in Redox reactions. F-actin is a specific substrate for these MICAL Redox enzymes, which oxidize specific amino acids within actin to destabilize actin filaments. Furthermore, this MICAL-catalyzed reaction is reversed by another family of Redox enzymes (SelR/MsrB enzymes)—thereby revealing a reversible Redox signaling process and biochemical mechanism regulating actin dynamics. Interestingly, in addition to the MICALs’ Redox enzymatic portion through which MICALs covalently modify and affect actin, MICALs have multiple other domains. Less is known about the roles of these other MICAL domains. Here we provide approaches for obtaining high levels of recombinant protein for the Redox only portion of Mical and demonstrate its catalytic and F-actin disassembly activity. These results provide a ground state for future work aimed at defining the role of the other domains of Mical — including characterizing their effects on Mical’s Redox enzymatic and F-actin disassembly activity.

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Role of hepcidin in oxidative stress and cell death of cultured mouse renal collecting duct cells: protection against iron and sensitization to cadmium

Archives of Toxicology ◽

10.1007/s00204-021-03106-z ◽

2021 ◽

Author(s):

Stephanie Probst ◽

Johannes Fels ◽

Bettina Scharner ◽

Natascha A. Wolff ◽

Eleni Roussa ◽

...

Keyword(s):

Oxidative Stress ◽

Cell Death ◽

Cell Fate ◽

Catalase Activity ◽

Metal Ion ◽

Iron Homeostasis ◽

Collecting Duct ◽

Duct Cell ◽

Plasmid Transfection

AbstractThe liver hormone hepcidin regulates systemic iron homeostasis. Hepcidin is also expressed by the kidney, but exclusively in distal nephron segments. Several studies suggest hepcidin protects against kidney damage involving Fe2+ overload. The nephrotoxic non-essential metal ion Cd2+ can displace Fe2+ from cellular biomolecules, causing oxidative stress and cell death. The role of hepcidin in Fe2+ and Cd2+ toxicity was assessed in mouse renal cortical [mCCD(cl.1)] and inner medullary [mIMCD3] collecting duct cell lines. Cells were exposed to equipotent Cd2+ (0.5–5 μmol/l) and/or Fe2+ (50–100 μmol/l) for 4–24 h. Hepcidin (Hamp1) was transiently silenced by RNAi or overexpressed by plasmid transfection. Hepcidin or catalase expression were evaluated by RT-PCR, qPCR, immunoblotting or immunofluorescence microscopy, and cell fate by MTT, apoptosis and necrosis assays. Reactive oxygen species (ROS) were detected using CellROX™ Green and catalase activity by fluorometry. Hepcidin upregulation protected against Fe2+-induced mIMCD3 cell death by increasing catalase activity and reducing ROS, but exacerbated Cd2+-induced catalase dysfunction, increasing ROS and cell death. Opposite effects were observed with Hamp1 siRNA. Similar to Hamp1 silencing, increased intracellular Fe2+ prevented Cd2+ damage, ROS formation and catalase disruption whereas chelation of intracellular Fe2+ with desferrioxamine augmented Cd2+ damage, corresponding to hepcidin upregulation. Comparable effects were observed in mCCD(cl.1) cells, indicating equivalent functions of renal hepcidin in different collecting duct segments. In conclusion, hepcidin likely binds Fe2+, but not Cd2+. Because Fe2+ and Cd2+ compete for functional binding sites in proteins, hepcidin affects their free metal ion pools and differentially impacts downstream processes and cell fate.

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Cellular and mitochondrial mechanisms of atrial fibrillation

Basic Research in Cardiology ◽

10.1007/s00395-020-00827-7 ◽

2020 ◽

Vol 115 (6) ◽

Author(s):

Fleur E. Mason ◽

Julius Ryan D. Pronto ◽

Khaled Alhussini ◽

Christoph Maack ◽

Niels Voigt

Keyword(s):

Atrial Fibrillation ◽

Nicotinamide Adenine Dinucleotide ◽

Molecular Mechanisms ◽

Treatment Options ◽

Specific Treatment ◽

Nicotinamide Adenine Dinucleotide Phosphate ◽

Nicotinamide Adenine ◽

Mitochondrial Activity ◽

Atp Production

AbstractThe molecular mechanisms underlying atrial fibrillation (AF), the most common form of arrhythmia, are poorly understood and therefore target-specific treatment options remain an unmet clinical need. Excitation–contraction coupling in cardiac myocytes requires high amounts of adenosine triphosphate (ATP), which is replenished by oxidative phosphorylation in mitochondria. Calcium (Ca2+) is a key regulator of mitochondrial function by stimulating the Krebs cycle, which produces nicotinamide adenine dinucleotide for ATP production at the electron transport chain and nicotinamide adenine dinucleotide phosphate for the elimination of reactive oxygen species (ROS). While it is now well established that mitochondrial dysfunction plays an important role in the pathophysiology of heart failure, this has been less investigated in atrial myocytes in AF. Considering the high prevalence of AF, investigating the role of mitochondria in this disease may guide the path towards new therapeutic targets. In this review, we discuss the importance of mitochondrial Ca2+ handling in regulating ATP production and mitochondrial ROS emission and how alterations, particularly in these aspects of mitochondrial activity, may play a role in AF. In addition to describing research advances, we highlight areas in which further studies are required to elucidate the role of mitochondria in AF.

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STUDIES ON THE SEEDLING DISEASE OF BARLEY CAUSED BY HELMINTHOSPORIUM SATIVUM P.K. & B.

Canadian Journal of Botany ◽

10.1139/b56-048 ◽

1956 ◽

Vol 34 (4) ◽

pp. 653-673 ◽

Cited By ~ 16

Author(s):

R. A. Ludwig ◽

R. V. Clark ◽

J. B. Julien ◽

D. B. Robinson

Keyword(s):

Disease Incidence ◽

Infested Soil ◽

Disease Development ◽

Toxic Activity ◽

Factors Affecting ◽

Seedling Disease ◽

Nutrient Salt ◽

Helminthosporium Sativum ◽

Treatment Comparisons

A standard sand – cornmeal – nutrient salt medium, for use in the production of artificial inoculum of Helminthosporium sativum, is described. This inoculum induces uniform plant disease development when thoroughly incorporated with the planting soil. The results presented clearly demonstrate the necessity of using a series of infestation levels in studies of factors affecting disease development in artificially infested soil. It is shown that considerable reliance can be placed on treatment comparisons within an experiment but that comparisons between experiments are much less accurate. The role of a toxin (or toxins) in disease development in barley seedlings has been demonstrated. The toxic activity was found to be distinct from that frequently encountered on addition of organic matter to soil. Results obtained suggest that toxin adsorption by the soil may play an important role in reducing disease incidence and severity.

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Akt isoforms differentially regulate neutrophil functions

Blood ◽

10.1182/blood-2009-11-255323 ◽

2010 ◽

Vol 115 (21) ◽

pp. 4237-4246 ◽

Cited By ~ 73

Author(s):

Jia Chen ◽

Haiyang Tang ◽

Nissim Hay ◽

Jingsong Xu ◽

Richard D. Ye

Keyword(s):

Kinase Inhibitor ◽

Nicotinamide Adenine Dinucleotide Phosphate ◽

Leading Edge ◽

Neutrophil Activation ◽

Enzyme Release ◽

Wild Type ◽

Akt Isoforms ◽

Phosphoinositide 3 Kinase ◽

O2 Production

In neutrophils, the phosphoinositide 3-kinase/Akt signaling cascade is involved in migration, degranulation, and O2− production. However, it is unclear whether the Akt kinase isoforms have distinct functions in neutrophil activation. Here we report functional differences between the 2 major Akt isoforms in neutrophil activation on the basis of studies in which we used individual Akt1 and Akt2 knockout mice. Akt2−/− neutrophils exhibited decreased cell migration, granule enzyme release, and O2− production compared with wild-type and Akt1−/− neutrophils. Surprisingly, Akt2 deficiency and pharmacologic inhibition of Akt also abrogated phorbol ester-induced O2− production, which was unaffected by treatment with the phosphoinositide 3-kinase inhibitor LY294002. The decreased O2− production in Akt2−/− neutrophils was accompanied by reduced p47phox phosphorylation and its membrane translocation, suggesting that Akt2 is important for the assembly of phagocyte nicotinamide adenine dinucleotide phosphate oxidase. In wild-type neutrophils, Akt2 but not Akt1 translocated to plasma membrane upon chemoattractant stimulation and to the leading edge in polarized neutrophils. In the absence of Akt2, chemoattractant-induced Akt protein phosphorylation was significantly reduced. These results demonstrate a predominant role of Akt2 in regulating neutrophil functions and provide evidence for differential activation of the 2 Akt isoforms in neutrophils.

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