Cor a 9

What is Cor a 9 and what is its biochemical classification?

Cor a 9, also known as corylin, belongs to the 11S globulin family of seed storage proteins within the cupin superfamily of proteins. It is classified as a dicupin, consisting of acidic and basic polypeptide chains bridged by disulfide linkages. The allergenic acidic polypeptide chain possesses a molecular weight of 40 kDa. Together with 7S albumins, these proteins constitute approximately 50% of the total seed storage proteins required for plant germination .

As a member of the 11S globulin family, Cor a 9 shares structural and functional similarities with other seed storage proteins. The protein demonstrates quaternary structure organization, typically forming a hexamer in its native state, as confirmed by size exclusion chromatography studies during purification processes .

What are the structural characteristics of Cor a 9 as determined by crystallographic studies?

Crystallographic studies have successfully obtained diffraction-quality single crystals of Cor a 9 using the hanging-drop vapor-diffusion method. X-ray diffraction data have been collected to 1.9 Å resolution using synchrotron radiation sources, allowing for detailed structural analysis .

The crystals belonged to the trigonal space group P3₁21 with unit-cell parameters a = b = 135.2 Å and c = 182.1 Å. When analyzed with the assumption of three Cor a 9 monomers in an asymmetric unit, the Matthews coefficient was estimated to be 3.39 ų/Da⁻¹, corresponding to protein crystals containing approximately 63.7% solvent . This high-resolution structural information provides essential insights for understanding epitope regions and protein-antibody interactions relevant to allergic responses.

What are the established protocols for extraction and purification of native Cor a 9?

The purification of native Cor a 9 involves a multi-step process that capitalizes on its biochemical properties:

• Initial Extraction: Hazelnut kernels are extracted using a high-salt solution (2M NaCl) at elevated temperature (333K, approximately 60°C) with periodic stirring, followed by centrifugation at 2000g for 15 minutes at room temperature .

• Defatting Process: The crude extract undergoes defatting through multiple cycles of hexane treatment (1:1 ratio) with 2-hour incubations at room temperature followed by centrifugation to separate the aqueous portion .

• Salt Precipitation: The salt concentration is reduced to 0.4M by dilution with water at 333K, causing Cor a 9 to precipitate during a 2-hour incubation at 273K (0°C). After centrifugation, the pellet is redissolved in 2M NaCl buffer .

• Chromatographic Purification: The sample undergoes gel filtration on a Superdex-200 column equilibrated with buffer containing 1M NaCl and 10mM Tris at pH 8.0. Fractions containing Cor a 9 are further purified using hydrophobic interaction chromatography on Phenyl HP columns with a decreasing ammonium sulfate gradient (2M to 0M) .

This methodology yields highly purified Cor a 9 suitable for crystallization and immunological studies, with protein concentration determined using its theoretical extinction coefficient (ε₂₈₀ = 47,900 cm⁻¹ mol⁻¹) .

What is the geographical distribution of Cor a 9 sensitization, and how does it vary across populations?

Sensitization to Cor a 9 exhibits notable geographical variations across Europe and North America:

The EuroPrevall study (2005-2010), which analyzed 731 hazelnut-sensitized individuals across 12 European centers, provides the most comprehensive epidemiological data on Cor a 9 sensitization patterns .

How does Cor a 9 sensitization correlate with clinical symptom severity?

Cor a 9 sensitization demonstrates a strong correlation with more severe clinical manifestations of hazelnut allergy:

• Symptom Severity Correlation: The EuroPrevall study showed a statistically significant association (p=0.025) between Cor a 9 sensitization and moderate-to-severe clinical presentations. Cor a 9 sensitization was present in 18.3% of patients with severe symptoms, 10.2% with moderate symptoms, and only 7.2% with oral allergy syndrome (OAS) .

• Anaphylaxis Association: In a study of 227 children with severe allergic reactions or anaphylaxis, Cor a 9 sensitization showed a 71.4% association with these serious manifestations .

• Systemic Reaction Pattern: A Belgian study (n=119) found Cor a 9 sensitization significantly more prevalent in patients with generalized reactions (erythema, urticaria, angioedema, gastrointestinal symptoms, and respiratory symptoms) compared to those with oral allergy syndrome (70% vs. 11%, p<0.05) .

• Respiratory Comorbidity: In 220 hazelnut-allergic asthmatic patients, those with positive specific IgE to Cor a 9 reported more generalized reactions, including gastrointestinal symptoms or anaphylaxis, compared to those without Cor a 9 sensitization .

These findings establish Cor a 9 as a clinically relevant marker for potentially severe hazelnut allergy manifestations, distinguishing it from pollen-related cross-reactive sensitization patterns.

What is the predictive value of Cor a 9-specific IgE in differentiating between sensitization and clinical allergy?

Cor a 9-specific IgE (sIgE) testing offers valuable diagnostic information but with certain limitations:

For optimal diagnostic accuracy, combining multiple component-resolved diagnostic markers (particularly Cor a 9 with Cor a 14) provides superior discriminatory power between clinically relevant allergy and asymptomatic sensitization.

How do various processing methods affect Cor a 9 allergenicity and stability?

Cor a 9 demonstrates considerable thermal and digestive resistance, though specific processing conditions can modulate its allergenicity:

• Thermal Processing Thresholds:

  • Roasting (hot air or infrared): Allergenicity remains stable up to 140°C

  • Infrared roasting at 170°C: Significant reduction in allergenicity

  • Autoclaving: Allergenicity affected by treatment at 121°C for 15 minutes or 138°C for 30 minutes

• Digestive Stability: Cor a 9 exhibits substantial resistance to digestive enzymes, contributing to its ability to trigger systemic allergic reactions affecting multiple organ systems .

• Structure-Function Relationship: The protein's inherent stability likely relates to its quaternary structure as a hexamer and the presence of disulfide bridges between acidic and basic polypeptide chains, providing resistance to denaturation under various processing conditions.

This stability profile explains why many hazelnut-allergic individuals react to both raw and conventionally processed hazelnuts, highlighting the importance of developing specialized processing methods for producing hypoallergenic hazelnut products.

What cross-reactivity exists between Cor a 9 and homologous proteins in other food sources?

Cross-reactivity studies reveal selective patterns of molecular relationships between Cor a 9 and other 11S globulins:

• Relationship with Peanut Allergens: Cor a 9 and peanut 11S globulin (Ara h 3) demonstrate limited cross-reactivity. Co-sensitization patterns show:

  • Virtually absent co-sensitization in hazelnut-allergic patients without peanut allergy

  • Approximately 10% co-sensitization in peanut-allergic patients without hazelnut allergy

  • Up to 30% co-sensitization in patients with both hazelnut and peanut allergies

• Mustard Seed Cross-Reactivity: The 11S globulins of mustard (Sin a 2) and hazelnut (Cor a 9) show significant cross-reactivity, as demonstrated in studies of mustard-allergic patients .

• Other Tree Nuts: While not explicitly detailed in the search results, the structural similarity between Cor a 9 and brazil nut 11S protein (Ber e 2) suggests potential cross-reactivity, with both proteins sharing physical and biological properties .

These cross-reactivity patterns have important implications for clinical management and allergen avoidance recommendations, particularly for patients with multiple nut allergies.

How does Cor a 9 sensitization vary between pediatric and adult populations?

Age-dependent variations in Cor a 9 sensitization profiles have been documented:

• Higher Pediatric Prevalence: Research demonstrates greater IgE reactivity toward Cor a 9 in children compared to adults. This age-related difference may reflect distinct sensitization mechanisms or exposures during developmental periods .

• Clinical Significance in Children: The association between Cor a 9 sensitization and severe allergic reactions appears particularly pronounced in pediatric populations. A study of 227 children with severe allergic reactions or anaphylaxis found a 71.4% association with Cor a 9 sensitization .

• Diagnostic Implications: The higher prevalence and potential clinical significance of Cor a 9 sensitization in children suggests that component-resolved diagnostics may have particular value for pediatric patients with suspected hazelnut allergy.

These age-related differences underscore the importance of age-appropriate clinical assessment and management strategies for hazelnut allergy across the lifespan.

What crystallization techniques have proven successful for structural studies of Cor a 9?

Successful crystallization of Cor a 9 has been achieved through careful optimization of several parameters:

• Concentration Screening: Crystallization trials systematically tested protein concentrations ranging from 21 to 170 mg/ml, with optimal results obtained at intermediate concentrations .

• Hanging-Drop Vapor Diffusion Method: Using EasyXtal CrystalSupports, 2 μl aliquots of protein solution were sealed over 0.5 ml reservoir solutions containing NaCl gradients (0-0.5M) at 0.05M intervals .

• Temperature Conditions: Parallel crystallization trials were conducted at both room temperature (298K) and 277K to identify optimal crystallization conditions .

• Cryoprotection Optimization: Various cryoprotectants (sucrose, glycerol, PEG 400, ethylene glycol) were screened, with 30% glycerol proving effective for preserving crystal integrity during flash-cooling in liquid nitrogen .

• Diffraction Analysis: High-quality diffraction data collection utilized synchrotron radiation at the SER-CAT 22-ID beamline (Advanced Photon Source), allowing resolution to 1.9 Å .

These methodological details provide valuable protocols for researchers aiming to reproduce or extend structural studies of Cor a 9 or related seed storage proteins.

What are the distinguishing features of Cor a 9-mediated sensitization compared to other hazelnut allergens?

Cor a 9 possesses two distinctive immunological characteristics that differentiate it from other hazelnut allergens:

• Primary Sensitization: Unlike pollen-related hazelnut allergens (e.g., Cor a 1) that often cause cross-reactive sensitization, Cor a 9 has the capacity to induce direct (primary) sensitization to hazelnut. This represents a fundamentally different mechanism of allergic sensitization, typically associated with stronger clinical reactivity .

• Severe Reaction Association: Cor a 9 sensitization correlates strongly with more severe systemic allergic manifestations, including skin reactions, respiratory symptoms, gastrointestinal involvement, and anaphylaxis. This contrasts with the predominantly mild, localized oral symptoms associated with pollen-related cross-sensitization .

Understanding these distinct mechanisms is crucial for accurate risk assessment and appropriate clinical management of hazelnut-allergic patients. The pattern of sensitization to specific molecular components can provide valuable predictive information about potential reaction severity and guide individualized treatment approaches.