Views: 3 Author: Jin Manhong, Ye Zhiwei, Zheng Qianwang, etc. Publish Time: 2022-12-14 Origin: Research progress on physiological activity, functional properties and application of edible mushroom protein[J]. China Food Additives, 2022 (12): 24-31.
Edible fungus is a traditional food with rich nutritional value, and scientists have been working to analyze its components with various functional properties in order to better serve human health. As one of the main components of edible fungi, protein accounts for about 19% to 37% of the dry weight. Oxidation and other functional activities. At present, the research on edible mushroom protein mainly focuses on the separation and functional identification of proteins with biological functions, and less attention is paid to its application in food processing. This article mainly reviews the current research progress on fungal proteins, including the physiological activity, digestibility, functional properties and processing applications of fungal proteins, as shown in Figure 1, in order to expand the use of fungal proteins in food processing. It provides a certain reference for its development and application.
1. Physiological activity of edible mushroom protein
Edible fungi contain a variety of bioactive proteins, mainly including Lectin, Fungal immunomodulatory proteins (FIPs), Ribosome-inactivating proteins (RIPs), Laccase, etc. Studies have shown that these proteins play important roles in anti-tumor, mitogenic, immune regulation, anti-virus, anti-inflammatory, anti-oxidation, etc., as shown in Table 1.
1.1. Lectins
Lectin is a non-immune protein or glycoprotein that can specifically bind to carbohydrates on the cell surface and has cell agglutination. Among all edible mushroom proteins, lectin is the most studied active protein. Agaricus bisporus lectin (ABL) can prevent DNA synthesis required for cell proliferation by binding to human colon epithelial cancer cell HT29-specific antigen galactose-β-1,3-N-acetylgalactosamine The protein enters the nucleus, thereby inhibiting cell proliferation; when the concentration of ABL is 25 μg/mL, it can achieve 87% inhibition [1-2]. Boletus edulis lectin (BEL) with homotetramer structure can show anti-proliferation effect on HT29 cells by recognizing N-acetylglucosamine residues (N-acetylglucosamine, GlcNAc) on HT29 cells, and in Low concentration (10μg/mL) can achieve a higher (92%) inhibitory effect [3].
Lectins exhibit mitogenic activity on lymphocytes. For example, Volvariella volvacea lectin (VVL) with a molecular weight of 12kDa can trigger cell signaling pathways to influx extracellular Ca2+ by binding to mouse spleen T lymphocyte receptors. And induce the expression of lymphocyte proliferation factor interleukin-2, thereby stimulating the proliferation of T lymphocytes [4]; VVL can also activate T cells and induce T lymphocyte proliferation by activating the signaling molecule tyrosine kinase p56 lck [5] . A sulfur bacteria lectin 4 (Laetiporus sulphureus letin 4, LSL 4) with a molecular weight of 76.0kDa can stimulate cells to secrete iNOS by recognizing Toll-like receptor 4 on the surface of macrophage RAW264. The effect of adjustment [6].
Lectins also exhibit certain antiviral activity against HIV virus and HBV virus. Lectins can display anti-HIV virus effects by recognizing the mannose-binding site on the surface of HIV virus, such as the 18.5kDa hygrophorus russula lectin composed of 175 amino acid residues, which can bind to Mannose-rich glycoprotein 120 binds, thereby inhibiting HIV virus from entering cells, or inhibiting HIV reverse transcriptase (HIV-RT) activity required for HIV virus proliferation to achieve anti-HIV virus effects [7]. Both Russula delica lectin (RDL) and Hericium erinaceum agglutinin (HEA) can inhibit the activity of HIV-RT, and RDL (half-inhibitory concentration IC50=0.26μM) exhibited a stronger effect than HEA (half-inhibition Concentration IC50 = 31.7μM) higher anti-HIV-RT activity [8-9]. Pleurotus ostreatus lectin (POL) can stimulate the production of HBV virus-specific antibodies by activating the Toll-like receptor 6 signaling pathway of dendritic cells, and finally achieve the effect of treating chronic HBV infection [10].
1.2. Immunomodulatory proteins
Fungal immunomodulatory proteins (FIPs) are a family of small molecular proteins with similar structures and sequences isolated from edible fungi, which have immunomodulatory activity. Fungal immunomodulatory proteins can play an immunomodulatory role in a variety of ways, including reducing the level of immunoglobulin E (IgE), affecting the immune function of T lymphocytes, and stimulating the secretion of cytokines. Ganoderma lucidum immunoregulatory protein (Ling Zhi-8) can inhibit the secretion of interleukin-4 by regulating helper T cells 2, thereby reducing the level of IgE associated with allergic reactions and inhibiting allergic reactions [11]. Lentinus tigrinus FIP-1/2, with a molecular weight of about 12kDa, can maintain the body’s Th1 and Th2 cells in a relatively balanced state by regulating the expression of transcription factors GATA-3 and T-bet, and achieve immune regulation effect [12]. The homodimeric structure of Trametes versicolor immunomodulatory protein prepared by ammonium sulfate precipitation can be used as an immunostimulator to promote macrophage RAW264.7 to secrete NO and tumor necrosis factor TNF-α to achieve the effect of immune stimulation [13]. Wang et al[14] carried out bioinformatics prediction and analysis on 7 kinds of immunoregulatory proteins in V. volvulus, and found that 7 kinds of FIPs consisted of 113 amino acid residues, with a molecular weight of about 13kDa, and all of them had immune activity; among them, Fip-vvo82 By inducing the release of interleukin-2, it regulates the immune response of the body and achieves the highest immunoregulatory activity, and its aliphatic index is the lowest, the protein instability index is the highest, and it contains higher content of valine and lysine. There is an obvious hydrophilic region between the 90th amino acid; the phylogenetic tree shows that Fip-vvo82 forms an independent lineage, and the other 6 species of V. volvulus FIPs are highly homologous.
Immunomodulatory proteins also have anti-tumor and anti-inflammatory activities. For example, recombinant Ganoderma lucidum immunomodulatory protein can induce apoptosis of lung cancer cell LLC1 and reduce tumor area in mice by down-regulating the expression level of heat shock protein [15]. The immunomodulatory protein of Flammulina velutipes with a molecular weight of about 13kDa can reduce the incidence of inflammation and respiratory diseases by inhibiting the replication of mouse respiratory syncytial virus and the expression of pro-inflammatory factor interleukin-6[16]. The immunomodulatory protein of Ganoderma microspore containing 111 amino acids can make microglial cells change from the classic activated state to the alternative activated state, and then release growth factor-β and interleukin-10, etc., so as to inhibit neuroinflammation and restore the balance of the body. Effect [17].
1.3. Ribosome-inactivating protein and laccase
Ribosome-inactivating proteins (RIPs) are enzymes that inactivate ribosomes by eliminating one or more adenosine residues in rRNA, and have the ability to inhibit ribosomes from synthesizing proteins. Current research focuses more on the anticancer physiological activity of edible fungus RIPs. The ribosome-inactivating protein Marmorin of Hypsizigus marmoreus can inhibit the proliferation of MCF7 breast cancer cells by inhibiting the expression of estrogen receptors, down-regulating the expression level of 17β-estradiol and reducing the binding between the two [18 ]. Another 20kDa ribosome inactivating protein, Hypsin, was isolated from the fruiting body of jiji mushroom, which can inhibit the proliferation of mouse leukemia cells and human leukemia and liver cancer cells [19]. Laccase is an oxidase that can oxidize polyphenols and bind multiple copper ions, and is closely related to the pathogenesis of tumors in organisms. Studies have shown that laccase can inhibit the replication of hepatitis C virus and the proliferation of leukemia cells (K562, Jurkat, HL-60) [20-21].
1.4. Other proteins and hydrolysates
A variety of mushroom proteins with antioxidant activity and their hydrolyzates have been reported successively, and it was also found that the molecular weight and amino acid composition of mushroom proteins are closely related to their antioxidant activity. Zheng Xinlei[22] isolated three kinds of proteins with different molecular weights from Pleurotus eryngii, and found that the DPPH free radical scavenging ability, superoxide anion free radical scavenging ability, Fe2+ It has the strongest chelating power and reducing ability. The peptides of Grifola frondosa fruiting body isolate protein hydrolyzed by trypsin showed strong antioxidant activity, among which the peptide fraction with the smallest molecular weight (<3kDa) showed the strongest antioxidant activity[23]. Similar results were also observed in the alkaline protease hydrolyzate of Schizophyllum commune protein, and the hydrolyzate with molecular weight less than 0.65kDa had the highest ability to scavenge ABTS free radicals[24]. Kimatu et al[25] compared the antioxidant activity of Agaricus bisporus protein isolate and its hydrolyzate, and found that the hydrolyzed polypeptide had higher content of negatively charged amino acids and hydrophobic amino acids (alanine, leucine, valine, etc.) than the protein isolate. amino acids); among them, negatively charged amino acids have abundant electrons that can be used to quench free radicals to enhance antioxidant activity, and hydrophobic amino acids can enhance the solubility of polypeptides in lipids to increase their interactions with free radicals or through hydrophobic associations. Incorporate into target organs to exert free radical scavenging ability.
2. Digestive properties of mushroom protein
The digestibility of protein is an important index to evaluate the nutritional value of protein. It refers to the percentage of protein absorbed by the human body after protein digestion, which is closely related to the difficulty of hydrolysis of peptide bonds. True protein digestibility (TPD) and in vitro protein digestibility (IVPD) can reflect the effect of protein digestion and absorption to a certain extent. Longvah［26］ and Dabbour［27］ found that the TPD of Lentinus edodes, Schizophyllum commune, Pleurotus otreatus and Agaricus macrosporus were 76.3%, 76.3%, 53.2%, 73.4%, 80.5%. Through in vitro digestion experiments, it was found that the IVPD of Pleurotus sajor-caju and Lentinus lepidus were 63.61% and 66.09% respectively［28］. The above results show that the protein digestibility of edible fungi is lower than that of casein (TPD=87.49%, IVPD=83.91%), which may be due to the fact that edible fungi contain more polyphenols and anti-nutritional factors, which affect the absorption and utilization of protein. The digestibility of edible mushroom protein can be improved to a certain extent by extracting isolated protein, mainly because some polyphenols and anti-nutritional factors can be removed during the extraction process. For example, the digestibility of Pleurotus ostreatus protein isolate is as high as 100% after in vitro gastrointestinal simulated digestion, which is much higher than that of Pleurotus ostreatus powder (23.5%) [29]. Treatment methods such as heating and ultrasound can improve the digestibility of edible mushroom protein by inactivating anti-nutritional factors, unfolding protein structure, and exposing enzyme cleavage sites. For example, heat treatment increased the IVPD of Agaricus abruptibulbus and Termitomyces globulus from 62.81% and 47.91% to 81.46% and 75.41%, respectively [30]. After the Pleurotus eryngii protein is heated (75°C, 30min) and ultrasonically treated (200W, 30min), the surface hydrophobicity of the protein is enhanced and the structure unfolds, exposing more protease action sites, thereby improving the IVPD of the protein［31-32 ].
3. Functional properties of edible mushroom protein
On the one hand, protein in food can provide nutrition for the human body; on the other hand, because of its unique functional properties (solubility, emulsification, foaming, etc.), it can be used as an ingredient in different types of food processing. In recent years, research on fungal proteins has begun to expand from the isolation and identification of biologically active proteins to the exploration of functional properties, laying a theoretical foundation for the application of fungal proteins in the food industry. Solubility is one of the most important functional properties of proteins, and other functional properties of proteins (such as emulsification, foaming) are closely related to solubility. Near the isoelectric point (pH3.5～5.0), the solubility of edible fungus protein is the lowest; at pH far away from the isoelectric point, the solubility gradually increases. Proteins isolated from various edible fungi have low solubility (<60%) under neutral conditions (pH7.0), such as black fungus protein [33], Coprinus comatus protein [34], Pleurotus eryngii protein [35], Flammulina velutipes protein mycorrhizal protein［36］, Bailing mushroom protein［37］ and white jade mushroom protein［38］, which largely limit its application in the food industry. In order to improve the dispersibility of edible fungus protein isolate in aqueous solution, our team adopted a protein granulation strategy to prepare edible fungus protein into nanoparticles that can be uniformly dispersed in aqueous solution. The albumin and total protein isolate of Bailing mushroom (Pleurotus tuoliensis) were prepared by adjusting the acid-base method to prepare nanoparticles with good dispersibility in aqueous solution, with an average particle size of 186.0-273.8nm and a potential of -26.94--29.78mV. The particle structure is mainly stabilized by hydrophobic interactions and disulfide bonds; these protein particles exhibit rapid interfacial adsorption at the oil-water interface and can be prepared with oil content ≤ 60%
stable emulsion [39]. Our team used the same method to prepare low-water-solubility boletus (Phlebopus portentosus) protein into nanoparticles. The three-phase contact angle of the particles at the oil-water interface is close to neutral (θo/w=95.62), which can be quickly adsorbed on The oil-water interface is assembled into an interface network structure in an orderly manner; interestingly, double emulsions can be prepared by a simple one-step homogenization method, which provides a new idea for the preparation of double emulsions[40]. It is worth noting that the solubility of albumin and globulin of Cordyceps militaris in neutral aqueous solution can be as high as 90%, which is much higher than that of other edible fungus isolate proteins; the hydrophobicity of albumin and globulin is significantly lower than that of gluten Protein, strong protein-interface hydrophobic interaction allows gluten to adsorb to the soda-water interface more quickly, showing better foaming properties, but the foam stability of albumin and globulin is significantly better than that of globulin[41] .
4. Application of edible mushroom protein in food processing
At present, some edible mushroom proteins have been used in food processing fields such as dairy products, beverages, baking, and artificial meat, as shown in Table 2. The enzyme extract isolated and purified from Pleurotus florida has the highest milk-clotting activity (367.85SU) at a temperature of 50°C and a pH of 6.0, and can be used as a substitute for calf rennet in cheese production［ 42]. The extracellular metalloprotease of Gallinarum gallinarum with a molecular weight of 29kDa has high milk coagulation activity (333.33U/mL) and low protein hydrolysis activity (5.21U/mL), and can also be used as curd for milk coagulation and cheese production Enzymes [43]. Immobilized laccase can be used as a beverage stabilizer. For example, treating white wine Moscato and Montonico with 1U/mL Yunzhi laccase can reduce the total phenolic content by 32.1% and 33.4% respectively, thereby preparing more stable wine[44]; The recombinant laccase POXA1b (2000U/g) prepared by immobilization of Pleurotus ostreatus can reduce the content of phenolic compounds in orange juice by 45%, and retain the flavanones with antioxidant activity[45]. Edible fungi can also be used as ingredients to add to baked products because of their high protein content, improving the quality of the product. Adding Agaricus bisporus powder (2.5%-7.5%) to black bread can increase the protein and vitamin D content, and the bread with 7.5% Agaricus bisporus powder has the highest overall acceptability. There was no significant difference in the mushroom powder control group［46］. Pteris mushroom powder (2%-6%) can be added to bread and cream cakes as a nutritional fortifier. When the addition amount is 6%, the protein content of bread and cake is as high as 10.01% and 12.33%, respectively, and the fat content is reduced to 1.99% and 13.89% [47]. Cookies are higher in carbohydrates, calories and fat, but lower in protein, vitamins and minerals. It has been reported that the content of protein and fiber in biscuits can be increased and the fat content can be reduced by adding anchovy powder and shiitake mushroom powder[48-49]. The fruiting bodies and mycelia of edible fungi are rich in protein and natural fiber structure, and have great potential in the preparation of meat substitutes. Button mushroom fruiting bodies were used to develop sausage analogues, and mushroom sausage analogues mixed with 0.8% carrageenan showed the best results in terms of textural properties, such as reduced storage and cooking losses, and improved emulsification Stability [50]. Our team used the fruiting bodies of Cordyceps militaris to prepare meatloaf substitutes in the previous work, and found that beating treatment can dissolve the protein in the fruiting bodies and leave a fibrous structure, so that the product forms a good fibrous structure and gel structure, thus endowing the product with high The firmness, chewiness, cohesion and sensory scores of the samples[51]. Compared with Russian animal sausage, sausage analogues made of oyster mushroom mycelium still had better hardness and crispness after storage at 2°C for four weeks[52]. Meat analogs prepared from Agaricus bisporus mycelium exhibited better elasticity and chewiness than soybean protein-based meat analogs, as well as more attractive umami characteristics[53].
5. Outlook
In order to meet the protein demand of population growth, people have begun to explore the functional properties of mushroom protein and its application in food. The rich protein components of edible fungi can not only serve as a good source of protein, but also endow it with physiological activities such as anti-tumor, immune regulation, anti-virus, and anti-oxidation. It can be expected that edible mushroom protein has great application potential in the food field. It can be used as a protein supplement to increase the protein content of the product. Physiologically active protein can also be prepared into various nutritional supplements and used in health food. However, the current research on edible fungal protein mainly focuses on protein extraction and functional activity identification, and there are few studies on the application of edible fungal protein in food. Therefore, we should pay more attention to and study the functional properties of fungal protein, develop it as a new protein resource and use it in food, and lay a theoretical basis for expanding the application of fungal protein in the food industry.
