The first-order reaction kinetics of the cryotrapped 1,1,2,2-
H
-aminoethanol substrate radical intermediate state in the adenosylcobalamin (B
)-dependent ethanolamine ammonia-lyase (EAL) from
...serovar Typhimurium are measured over the range of 203-225 K by using time-resolved, full-spectrum electron paramagnetic resonance spectroscopy. The studies target the fundamental understanding of the mechanism of EAL, the signature enzyme in ethanolamine utilization metabolism associated with microbiome homeostasis and disease conditions in the human gut. Incorporation of
H into the hydrogen transfer that follows the substrate radical rearrangement step in the substrate radical decay reaction sequence leads to an observed
H/
H isotope effect of approximately 2 that preserves, with high fidelity, the idiosyncratic piecewise pattern of rate constant versus inverse temperature dependence that was previously reported for the
H-labeled substrate, including a monoexponential regime (
≥ 220 K) and two distinct biexponential regimes (
= 203-219 K). In the global kinetic model, reaction at ≥220 K proceeds from the substrate radical macrostate,
, and at 203-219 K along parallel pathways from the two sequential microstates,
and
, that are distinguished by different protein configurations. Decay from
, or
and
, is rate-determined by radical rearrangement (
H) or by contributions from both radical rearrangement and hydrogen transfer (
H). Non-native direct decay to products from
is a consequence of the free energy barrier to the native
→
protein configurational transition. At physiological temperatures, this is averted by the fast protein configurational dynamics that guide the
→
transition.
Full text
Available for:
IJS, KILJ, NUK, PNG, UL, UM
Some prokaryotes compartmentalize select metabolic capabilities. Salmonella enterica subspecies enterica serovar Typhimurium LT2 (hereafter S. Typhimurium) catabolizes ethanolamine (EA) within a ...proteinaceous compartment that we refer to as the ethanolamine utilization (Eut) metabolosome. EA catabolism is initiated by the adenosylcobalamin (AdoCbl)‐dependent ethanolamine ammonia‐lyase (EAL), which deaminates EA via an adenosyl radical mechanism to yield acetaldehyde plus ammonia. This adenosyl radical can be quenched, requiring the replacement of AdoCbl by the ATP‐dependent EutA reactivase. During growth on ethanolamine, S. Typhimurium synthesizes AdoCbl from cobalamin (Cbl) using the ATP:Co(I)rrinoid adenosyltransferase (ACAT) EutT. It is known that EAL localizes to the metabolosome, however, prior to this work, it was unclear where EutA and EutT localized, and whether they interacted with EAL. Here, we provide evidence that EAL, EutA, and EutT localize to the Eut metabolosome, and that EutA interacts directly with EAL. We did not observe interactions between EutT and EAL nor between EutT and the EutA/EAL complex. However, growth phenotypes of a ΔeutT mutant strain show that EutT is critical for efficient ethanolamine catabolism. This work provides a preliminary understanding of the dynamics of AdoCbl synthesis and its uses within the Eut metabolosome.
Ethanolamine ammonia‐lyase (EAL) uses coenzyme B12 (CoB12) to catabolize the first step of the ethanolamine catabolic pathway. If CoB12 is oxidized, EAL becomes inactive. We show in vitro and in vivo evidence that the EutA reactivase directly interacts with EAL inside the metabolosome, where ethanolamine catabolism takes place. The EutT enzyme that synthesizes CoB12 also interacts with the metabolosome, but not directly with the EutA‐EAL complex.
Full text
Available for:
BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
In recent decades, nanopores have become a promising diagnostic tool. Protein and solid-state nanopores are increasingly used for both RNA/DNA sequencing and small molecule detection. The latter is ...of great importance, as their detection is difficult or expensive using available methods such as HPLC or LC-MS. DNA aptamers are an excellent detection element for sensitive and specific detection of small molecules. Herein, a method for quantifying small molecules using a ready-to-use sequencing platform is described. Taking ethanolamine as an example, a strand displacement assay is developed in which the target-binding aptamer is displaced from the surface of magnetic particles by ethanolamine. Non-displaced aptamer and thus the ethanolamine concentration are detected by the nanopore system and can be quantified in the micromolar range using our in-house developed analysis software. This method is thus the first to describe a label-free approach for the detection of small molecules in a protein nanopore system.
Full text
Available for:
GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Glycerophospholipids including phosphatidylethanolamine (PE) and phosphatidylcholine (PC) are vital components of biological membranes. Trypanosomatid parasites of the genus Leishmania can acquire PE ...and PC via de novo synthesis and the uptake/remodeling of host lipids. In this study, we investigated the ethanolaminephosphate cytidylyltransferase (EPCT) in Leishmania major, which is the causative agent for cutaneous leishmaniasis. EPCT is a key enzyme in the ethanolamine branch of the Kennedy pathway which is responsible for the de novo synthesis of PE. Our results demonstrate that L. major EPCT is a cytosolic protein capable of catalyzing the formation of CDP-ethanolamine from ethanolamine-phosphate and cytidine triphosphate. Genetic manipulation experiments indicate that EPCT is essential in both the promastigote and amastigote stages of L. major as the chromosomal null mutants cannot survive without the episomal expression of EPCT. This differs from our previous findings on the choline branch of the Kennedy pathway (responsible for PC synthesis) which is required only in promastigotes but not amastigotes. While episomal EPCT expression does not affect promastigote proliferation under normal conditions, it leads to reduced production of ethanolamine plasmalogen or plasmenylethanolamine, the dominant PE subtype in Leishmania. In addition, parasites with episomal EPCT exhibit heightened sensitivity to acidic pH and starvation stress, and significant reduction in virulence. In summary, our investigation demonstrates that proper regulation of EPCT expression is crucial for PE synthesis, stress response, and survival of Leishmania parasites throughout their life cycle.
Full text
Available for:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
The interfacial dipolar polarization in inverted structure polymer solar cells, which arises spontaneously from the absorption of ethanolamine end groups, such as amine and hydroxyl groups on ...ripple‐structure zinc oxide (ZnO‐R), lowers the contact barrier for electron transport and extraction and leads to enhanced electron mobility, suppression of bimolecular recombination, reduction of the contact resistance and series resistance, and remarkable enhancement of the power conversion efficiency.
Full text
Available for:
BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
The Warburg effect is the major metabolic hallmark of cancer. According to Warburg himself, the consequence of the Warburg effect is cell dedifferentiation. Therefore, reversing the Warburg effect ...might be an approach to restore cell differentiation in cancer. In this study, we used a mitochondrial uncoupler, niclosamide ethanolamine (NEN), to activate mitochondrial respiration, which induced neural differentiation in neuroblastoma cells. NEN treatment increased the NAD+/NADH and pyruvate/lactate ratios and also the α-ketoglutarate/2-hydroxyglutarate (2-HG) ratio. Consequently, NEN treatment induced promoter CpG island demethylation and epigenetic landscape remodeling, activating the neural differentiation program. In addition, NEN treatment upregulated p53 but downregulated N-Myc and β-catenin signaling in neuroblastoma cells. Importantly, even under hypoxia, NEN treatment remained effective in inhibiting 2-HG generation, promoting DNA demethylation, and suppressing hypoxia-inducible factor signaling. Dietary NEN intervention reduced tumor growth rate, 2-HG levels, and expression of N-Myc and β-catenin in tumors in an orthotopic neuroblastoma mouse model. Integrative analysis indicated that NEN treatment upregulated favorable prognosis genes and downregulated unfavorable prognosis genes, which were defined using multiple neuroblastoma patient datasets. Altogether, these results suggest that mitochondrial uncoupling is an effective metabolic and epigenetic therapy for reversing the Warburg effect and inducing differentiation in neuroblastoma.
Targeting cancer metabolism using the mitochondrial uncoupler niclosamide ethanolamine leads to methylome reprogramming and differentiation in neuroblastoma, providing a therapeutic opportunity to reverse the Warburg effect and suppress tumor growth. See related commentary by Byrne and Bell, p.167.
Unhealthy dietary habits have been identified as a risk factor for the development and progression of cancer. Therefore, adopting a healthy eating pattern is currently recommended to prevent the ...onset of different types of cancers, including breast carcinoma. In particular, the Mediterranean diet, based on high consumption of omega-3 polyunsaturated fatty acids (
-3 PUFAs), such as those found in cold-water fish and other seafood, nuts, and seeds, is recommended to reduce the incidence of several chronic-degenerative diseases. Indeed, the consumption of
-3 PUFAs, particularly eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), reduced the risk of different types of cancer, including breast cancer. Moreover, they can counteract breast cancer progression and reduce the side effects of chemotherapy in breast cancer survival. Studies have demonstrated that DHA, exhibiting greater antitumor activity than EPA in breast cancer, can be attributed to its direct impact on breast cancer cells and also due to its conversion into various metabolites.
-docosahexaenoyl ethanolamine, DHEA, is the most studied DHA derivative for its therapeutic potential in breast cancer. In this review, we emphasize the significance of dietary habits and the consumption of
-3 polyunsaturated fatty acids, particularly DHA, and we describe the current knowledge on the antitumoral action of DHA and its derivative DHEA in the treatment of breast cancer.
Full text
Available for:
IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
N‐acylethanolamines, which include the endocannabinoid anandamide and the cannabinoid receptor‐inactive saturated compounds N‐palmitoyl ethanolamine and N‐stearoyl ethanolamine, are ethanolamines of ...long‐chain fatty acids degraded by fatty acid amide hydrolase (FAAH) known to accumulate in degenerating tissues and cells. Whilst much evidence supports a protective anti‐inflammatory role of both anandamide and N‐palmitoyl ethanolamine, very little information is available with regard to the bioactivity of N‐stearoyl ethanolamine. Employing a murine model of passive IgE‐induced cutaneous anaphylaxis, we have found that N‐stearoyl ethanolamine is endowed with marked anti‐inflammatory properties in vivo, supporting the hypothesis that endogenous N‐stearoyl ethanolamine is, in analogy to N‐palmitoyl ethanolamine, a bioactive signalling lipid capable of downregulating allergic inflammation in the skin. This effect, although mimicked by synthetic, non‐selective, CB1/CB2 receptor agonists, such as WIN55, 212‐2, was not sensitive to CB1 or CB2 receptor antagonists, but rather was fully reversed by capsazepine, a competitive antagonist of the TRPV1 receptor. Moreover, CB1 receptor antagonists, although effective in antagonising the WIN55,212‐2‐induced hypothermia, did not reduce the anti‐inflammatory effect of WIN55,212‐2, whilst CB2 receptor antagonists, per se inactive, potentiated the WIN55,212‐2 effect, suggesting an involvement of non‐CB1/CB2 receptors in the anti‐inflammatory action of WIN55,212‐2. All this, together with demonstration of FAAH as a major regulator of the in vivo concentrations of saturated N‐stearoyl ethanolamine, in addition to N‐palmitoyl ethanolamine, raise the speculation that pharmacological treatments with saturated N‐acylethanolamines such as N‐stearoyl ethanolamine, or alternatively FAAH inhibitors able to increase their local concentration, rather than selective CB receptor agonists, might be of promising therapeutic benefit in reducing allergic inflammation in the skin.
Full text
Available for:
BFBNIB, DOBA, FZAB, GIS, IJS, IZUM, KILJ, NLZOH, NUK, OILJ, PILJ, PNG, SAZU, SBCE, SBMB, UILJ, UKNU, UL, UM, UPUK
Phospholipids are the primary constituents of cell membranes across all domains of life, but how and when phospholipids appeared on early Earth remains unknown. Pressingly, most prebiotic syntheses ...of complex phospholipids rely upon substrates not yet shown to have been available on early Earth. Here, we describe potentially prebiotic syntheses of a diverse array of complex phospholipids and their building blocks. First, we show that choline could have been produced on early Earth by stepwise
-methylation of ethanolamine. Second, taking a systems chemistry approach, we demonstrate that the intrinsically activated glycerol-2,3-cyclic phosphate undergoes ring opening with combinations of prebiotic amino alcohols to yield complex phospholipid headgroups. Importantly, this pathway selects for the formation of 2-amino alcohol-bearing phospholipid headgroups and enables the accumulation of their natural regioisomers. Finally, we show that the dry-state ring opening of cyclic lysophosphatidic acids leads to a range of self-assembling lysophospholipids. Our results provide new prebiotic routes to key intermediates on the way toward modern phospholipids and illuminate the potential origin and evolution of cell membranes.
Full text
Available for:
IJS, KILJ, NUK, PNG, UL, UM
PDF
