Recent studies revealed that Amblyomma or Ixodes tick bites may cause red meat allergy, in which galactose‐α‐1,3‐galactose (α‐Gal) is a major IgE‐binding epitope. The incidence of red meat allergy is ...high in Shimane Prefecture, as is tick‐transmitted Japanese spotted fever. Therefore, we speculated that tick bites may cause these meat allergies. The carbohydrate α‐Gal was detected in the salivary gland protein of Haemaphysalis longicornis (H. longicornis), the vector for Japanese spotted fever, by immunoblotting using anti‐α‐Gal antibody. H. longicornis salivary gland protein‐specific IgE was detected in the sera of 24 of 30 patients with red meat allergies. Sensitization to tick salivary gland protein containing α‐Gal is possibly a major etiology of red meat allergy; the carbohydrate plays a crucial role in its allergenicity. These results further indicate that the α‐Gal epitope is present not only in Amblyomma or Ixodes, but also in Haemaphysalis.
Background
The production of IgE molecules specific to the carbohydrate galactose‐α‐1,3‐galactose (alpha‐gal) is known to induce delayed anaphylaxis against mammalian meat. Tick bites constitute the ...primary sensitization source, as ticks transfer alpha‐gal in their saliva to a host during a bite. The reported prevalence of alpha‐gal‐specific IgE (alpha‐gal‐sIgE) positivity varies between different populations from diverse geographic regions.
Objective
To investigate the prevalence of alpha‐gal‐sIgE positivity in a population of forest service employees who are highly exposed to ticks in comparison with a residential population and a historic sample.
Methods
A cross‐sectional study evaluating 300 forest service employees and hunters from southwest Germany was performed. Alpha‐gal‐sIgE levels were assessed by ImmunoCAP assay. The prevalence of alpha‐gal‐sIgE‐positive individuals was compared with a matched cohort composed of a residential population and blood samples from forest service employees collected 15 years ago.
Results
In the study population, the prevalence of alpha‐gal‐sIgE‐positive (≥0.10 kUA/L) individuals was 35.0%, whereas the prevalence of individuals with alpha‐gal‐sIgE levels ≥0.35 kUA/L was 19.3%. Alpha‐gal‐sIgE positivity was associated with total IgE levels and recent tick bites. Mammalian meat‐induced delayed anaphylaxis was found in 8.6% of the participants with alpha‐gal‐sIgE levels ≥0.35 kUA/L. For forest service employees and hunters, the odds ratio for alpha‐gal‐sIgE positivity was 2.48 compared to the residential population. The prevalence of alpha‐gal‐sIgE positivity in the current and historic cohort was comparable.
Conclusion
Forest service employees and hunters compose a population with a high prevalence of alpha‐gal‐sIgE positivity and carry a considerable risk of red meat allergy.
SUMMARY
Ascorbate plays an indispensable role in plants, functioning as both an antioxidant and a cellular redox buffer. It is widely acknowledged that the ascorbate biosynthesis in the ...photosynthetic tissues of land plants is governed by light‐mediated regulation of the D‐mannose/L‐galactose (D‐Man/L‐Gal) pathway. At the core of this light‐dependent regulation lies the VTC2 gene, encoding the rate‐limiting enzyme GDP‐L‐Gal phosphorylase. The VTC2 expression is regulated by signals via the photosynthetic electron transport system. In this study, we directed our attention to the liverwort Marchantia polymorpha, representing one of the basal land plants, enabling us to conduct an in‐depth analysis of its ascorbate biosynthesis. The M. polymorpha genome harbors a solitary gene for each enzyme involved in the D‐Man/L‐Gal pathway, including VTC2, along with three lactonase orthologs, which may be involved in the alternative ascorbate biosynthesis pathway. Through supplementation experiments with potential precursors, we observed that only L‐Gal exhibited effectiveness in ascorbate biosynthesis. Furthermore, the generation of VTC2‐deficient mutants through genome editing unveiled the inability of thallus regeneration in the absence of L‐Gal supplementation, thereby revealing the importance of the D‐Man/L‐Gal pathway in ascorbate biosynthesis within M. polymorpha. Interestingly, gene expression analyses unveiled a distinct characteristic of M. polymorpha, where none of the genes associated with the D‐Man/L‐Gal pathway, including VTC2, showed upregulation in response to light, unlike other known land plants. This study sheds light on the exceptional nature of M. polymorpha as a land plant that has evolved distinctive mechanisms concerning ascorbate biosynthesis and its regulation.
Significance Statement
Ascorbate biosynthesis in the liverwort M. polymorpha functions through the known D‐mannose/L‐galactose pathway, but unlike existing plants and green algae, the expression of the VTC2 gene encoding its key enzyme is completely devoid of any regulatory mechanism by light or reactive oxygen species. Our findings posit M. polymorpha as a terrestrial model plant of scholarly significance, providing invaluable insights into the evolutionary foundations of ascorbate biosynthesis and its regulation mechanisms.
The synthesis and reactivity in an 4+2 inverse electron demand hetero Diels‐Alder reaction (ihDA) of an original class of electron‐poor heterodienes, the ...N‐substituted‐1H‐benzoc1,2thiazin‐4‐one‐2,2‐dioxides, are described. These are highly reactive electrophiles that allow easy access to unprecedented benzo‐thiazine glyco‐fused derivatives in a remarkably selective way, even when using acetylated glycals, previously unexplored within this version of ihDA. DFT calculations support the experimental data, and moreover show that acetylated dienophiles can easily react making cycloadditions feasible.In memory of Prof. Giovanni Romeo, who gave a fundamental contribution to the field of cycloaddition reactions
3,6-Anhydro-
l
-galactose (L-AHG) constitutes 50 % of agarose, which is the main component of red macroalgae. No information is currently available on the mass production, metabolic fate, or ...physiological effects of L-AHG. Here, agarose was converted to L-AHG in the following three steps: pre-hydrolysis of agarose into agaro-oligosaccharides by using acetic acid, hydrolysis of the agaro-oligosaccharides into neoagarobiose by an exo-agarase, and hydrolysis of neoagarobiose into L-AHG and galactose by a neoagarobiose hydrolase. After these three steps, L-AHG was purified by adsorption and gel permeation chromatographies. The final product obtained was 95.6 % pure L-AHG at a final yield of 4.0 % based on the initial agarose. In a cell proliferation assay, L-AHG at a concentration of 100 or 200 μg/ mL did not exhibit any significant cytotoxicity. In a skin whitening assay, 100 μg/ mL of L-AHG showed significantly lower melanin production compared to arbutin. L-AHG at 100 and 200 μg/ mL showed strong anti-inflammatory activity, indicating the significant suppression of nitrite production. This is the first report on the production of high-purity L-AHG and its physiological activities.
Aging, a time‐dependent multifaceted process, affects both cell structure and function and involves oxidative stress as well as glycation. The present investigation focuses on the role of the band 3 ...protein (B3p), an anion exchanger essential to red cells homeostasis, in a d‐galactose (
d‐Gal)‐induced aging model. Anion exchange capability, measured by the rate constant of SO₄²− uptake through B3p, levels of lipid peroxidation, oxidation of membrane sulfhydryl groups, B3p expression, methemoglobin, glycated hemoglobin (Hb), and the reduced glutathione/oxidized glutathione ratio were determined after exposure of human erythrocytes to 25, 35, 50, and 100 mmol/L d‐Gal for 24 h. Our results show that: (i) in vitro application of d‐Gal is useful to model early aging in human erythrocytes; (ii) assessment of B3p ion transport function is a sensitive tool to monitor aging development; (iii) d‐Gal leads to Hb glycation and produces substantial changes on the endogenous antioxidant system; (iv) the impact of aging on B3p function proceeds through steps, first involving Hb glycation and then oxidative events at the membrane level. These findings offer a useful tool to understand the mechanisms of aging in human erythrocytes and propose B3p as a possible target for new therapeutic strategies to counteract age‐related disturbances.
Fungal copper radical oxidases (CROs) from the Auxiliary Activity family 5 (AA5) constitute a group of metalloenzymes that oxidize a wide panel of natural compounds, such as galactose‐containing ...saccharides or primary alcohols, into product derivatives exhibiting promising biotechnological interests. Despite a well‐conserved first copper‐coordination sphere and overall fold, some members of the AA5_2 subfamily are incapable of oxidizing galactose and galactosides but conversely efficiently catalyse the oxidation of diverse aliphatic alcohols. The objective of this study was to understand which residues dictate the substrate preferences between alcohol oxidases and galactose oxidases within the AA5_2 subfamily. Based on structural differences and molecular modelling predictions between the alcohol oxidase from Colletotrichum graminicola (CgrAlcOx) and the archetypal galactose oxidase from Fusarium graminearum (FgrGalOx), a rational mutagenesis approach was developed to target regions or residues potentially driving the substrate specificity of these enzymes. A set of 21 single and multiple CgrAlcOx variants was produced and characterized leading to the identification of six residues (W39, F138, M173, F174, T246, L302), in the vicinity of the active site, crucial for substrate recognition. Two multiple CgrAlcOx variants, i.e. M4F (W39F, F138W, M173R and T246Q) and M6 (W39F, F138W, M173R, F174Y, T246Q and L302P), exhibited a similar affinity for carbohydrate substrates when compared to FgrGalOx. In conclusion, using a rational site‐directed mutagenesis approach, we identified key residues involved in the substrate selectivity of AA5_2 enzymes towards galactose‐containing saccharides.
Fungal copper radical oxidases (CROs) from the AA5_2 subfamily can oxidize a wide panel of natural compounds including galactose‐containing saccharides or primary alcohols, but the molecular determinants driving the substrate specificity of these metalloenzymes were not clearly established. Using a rational site‐directed mutagenesis approach based on molecular modelling predictions, we identified several key amino acids driving affinity towards galactose‐containing substrates.
Patients with IgE antibodies against the carbohydrate epitope galactose‐α‐1,3‐galactose (α‐Gal) have reported severe allergic reactions after consumption of red meat. Investigations have revealed ...associations between IgE to α‐Gal and tick bites. We provide the first direct evidence that α‐Gal is present within ticks thus potentially explaining the relationship between tick exposure and sensitization to α‐Gal, with development of red meat allergy as a secondary phenomena. Serum from Swedish patients with delayed severe reactions to red meat was included in the study. A dose‐dependent inhibition of IgE responses to α‐Gal by the tick Ixodes ricinus is demonstrated. Furthermore, using cryostat‐cut sections of I. ricinus, we show that both a monoclonal and a polyclonal antibody against α‐Gal stains the gastrointestinal tract of the tick. The same pattern is seen when staining with patient sera IgE positive to α‐Gal. These results confirm that the α‐Gal epitope is present in I. ricinus and imply host exposure to α‐Gal during a tick bite. This provides further evidence that tick bites are associated with IgE responses to α‐Gal and red meat allergy.
Background
Serum IgE antibodies directed at galactose‐α‐1,3‐galactose (α‐Gal) are associated with a novel form of delayed anaphylaxis occurring upon consumption of red meat or innards. Pork kidney is ...known as the most potent trigger of this syndrome, but the culprit allergens have not yet been identified. The aim of this study was the identification and characterization of pork kidney proteins mediating delayed anaphylactic reactions through specific IgE to α‐Gal.
Methods
A cohort of 59 patients with specific IgE to α‐Gal was screened by immunoblot for IgE‐reactive proteins in pork kidney. Proteins were identified by peptide mass fingerprinting. Isolated proteins were assayed in ELISA and ELISA inhibition, basophil activation and skin prick test.
Results
Several IgE‐binding proteins of high molecular weight (100– >200 kDa) were detected in pork kidney extracts by immunoblot using patient sera and an anti‐α‐Gal antibody. Two major IgE‐binding proteins were identified as porcine angiotensin‐I‐converting enzyme (ACE I) and aminopeptidase N (AP‐N). Reactivity of patient sera and anti‐α‐Gal antibody to both proteins was abolished by carbohydrate oxidation. The α‐Gal IgE epitopes were resistant to heat denaturation. Pork kidney extract, isolated ACE I, and AP‐N were able to activate patient basophils and elicit positive responses in skin prick tests.
Conclusion
Two cell‐membrane proteins carrying α‐Gal epitopes were identified in pork kidney. For the first time, isolated meat proteins were shown to induce basophil activation in patients with delayed anaphylaxis to red meat providing further confirmation for the clinical relevance of these α‐Gal‐carrying proteins.