23, 397–402. (1992). The editor and reviewers' affiliations are the latest provided on their Loop research profiles and may not reflect their situation at the time of review. In the pre-screening stage, the morphology of the colony and the ratio of the clarification halo by protease activity to the colony diameter were the predominant factors used for selection. Microbiol. This fungi is essential to the fermentation processes because of its ability to secrete large amounts of various degrading enzymes, which allows it to decom… PubMed:Transformation of a serine protease of Aspergillus oryzae into a thiol-enzyme. Appl. The untreated samples served as the control of 100% survival rate. Branching mutants of Aspergillus oryzae with improved amylase and protease production on solid substrates. In addition, various bacterial species such as Bacillus (Bacillus amyloliquefaciens, B. subtilis, and B. natto) and lactic acid bacteria (LAB), including Tetragenococcus halophilus have been previously used [8, 10-12]. The A. oryzae strain 3.042 was purchased from China General Microbial Culture Collection Center (CGMCC No. After ARTP treatment, the isolation and screening of target mutants are key procedures in microbial breeding. (2005). Niu, F. X., He, X., Wu, Y. Q., and Liu, J. Beijing: General Administration of Quality Supervision. Simultaneous quantification of twenty Amadori products in soy sauce using liquid chromatography-tandem mass spectrometry. On le considère comme un moût, un malt ou un levain « d’amorçage » et il s’obtient par transformation de l’amidon présent dans les graines par l’Aspergillus oryzae. Int. Gene regulation in Aspergillus oryzae promotes hyphal growth and flavor formation in soy sauce koji. Appl. Agric. doi: 10.1271/bbb.80500, Lowry, O. H., Rosebrough, N. J., Farr, A. L., and Randall, R. J. 18:30. doi: 10.1186/s12934-019-1084-2, Zhang, W., Si, G., Rao, Z., Li, J., Zhang, X., Mei, J., et al. 42, 575–582. (2019). Biot. Eur. Appl. (2019). The purification and rejuvenation of Aspergillus oryzae Shanghai brew 3.042. Appl. XY helped to conduct the ARTP mutagenesis with WM and JS. 26, 352–355. Wang, X. L., Li, Z. W., Chen, L. H., Liu, X. Soy sauce production by solid-state fermentation involves many proteolytic and carbohydrase enzymes. A 10-μL spore suspension was spread over an iron dish and then air-dried under aseptic conditions. Aspergillus (A. oryzae, A. sojae, and A. awamori), Actinomucor taiwanensis, Rhizopus, and yeast (Zygosaccharomyces rouxii, and Saccharomyces cerevisiae). Furthermore, the strain B-2 exhibited increases of 54.7 and 17.3% in acid protease and neutral protease activity, respectively, whereas the activity of alkaline protease showed a decrease of 8.1% as compared with the original strain 3.042 (Figure 6). 102, 8739–8751. Comparative evaluation of neutral protease production by Aspergillus oryzae in submerged and solid-state fermentation. 164, 148–154. Microbiol. (2008). Casein and Folin-Ciocalteu’s phenol reagent were purchased from Beijing Solarbio Science and Technology Co., Ltd. (Beijing, China). Naturally fermented broad beans are the main raw material in Sichuan broad-bean sauce. (A) Ratio of the clarification halo to the colony diameter. doi: 10.1007/s00253-018-8937-1, Keywords: atmospheric and room temperature plasma, Aspergillus oryzae, protease, mutation, breeding, fermentation, Citation: Shu L, Si X, Yang X, Ma W, Sun J, Zhang J, Xue X, Wang D and Gao Q (2020) Enhancement of Acid Protease Activity of Aspergillus oryzae Using Atmospheric and Room Temperature Plasma. Probably is the signaling protease that mediates the first proteolytic cleavage within the signaling protease box of pacC (By similarity). Aspergillus oryzae (syn. (2014). Figure 1. Strain B-2 (CGMCC No. In particular, lactic acid buffer (0.05 mol/L, pH 3.0), phosphate buffer (0.02 mol/L, pH 7.2) and sodium tetraborate-NaOH buffer (0.0125 mol/L sodium tetraborate, 0.043 mol/L NaOH, pH 10.0) were used for the crude extraction of acid, neutral, and alkaline protease activity, respectively. doi: 10.1007/s00253-005-1968-4, Wang, L. Y., Huang, Z. L., Li, G., Zhao, H. X., Xing, X. H., Sun, W. T., et al. J. (2017). 5, 24224–24230. oryzae) est un champignon microscopique, de la classe des ascomycètes, appartenant aux moisissures « nobles », proche de Penicillium, et donc des moisissures des fromages à pâte persillée. Biosci. Compared to the “traditional” techniques, ARTP can generate diversified damage mechanism to genetic material, which resulting in the diversity of mutant types. Figure 5. Production of acid protease using thin stillage from a rice spirit distillery by Aspergillus niger. 69, 44–50. doi: 10.1111/j.1365-2621.2011.02870.x, Zhao, G., Yao, Y., Hao, G., Fang, D., Yin, B., Cao, X., et al. Cell Fact. Food Res. Of the 19 examined strains, 5 mutants, namely A-1, B-1, B-2, B-8, and B-13, exhibited much higher acid protease activities compared to the original strain 3.042 by increases of 58.1, 20.3, 39.9, 30.4, and 26.4%, respectively (Figure 4B). Evolutionary engineering of industrial microorganisms-strategies and applications. (2008) have identified the mechanism of radio-frequency and atmospheric-pressure glow discharges (APGD) directly acted on plasmid DNA and oligonucleotides. Genome sequencing and analysis of Aspergillus oryzae. Functional effects of Japanese style fermented soy sauce (shoyu) and its components. J. Biosci. Food Chem. Fermentation time, inoculum size, initial moisture content, salt solutions, and the thickness of the substrate were investigated one by one. However, it remains unclear whether there is an essential relationship between acid protease activity and colony morphology. Originally, A. oryzae strain 3.042 was obtained from a wild-type strain via multiple UV mutation, and the acid protease activity improvement has become more and more difficult. Int. Rev. J. Biol. The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. Front. Soy sauce, a traditional condiment in China, Japan, South Korea, and Southeast Asia, is fermented from soybean and wheat flour and is now used in many countries worldwide. 3.00951, Beijing, China) and used as the original strain for ARTP mutagenesis. In addition, this organism is also widely used for the manufacturing of various traditionally fermented foods (Zhao et al., 2013; Kitamato, 2015). Une large gamme d'options de aspergillus oryzae s'offre à vous comme des réagents généraux, des nématoïde et des agent auxiliaire chimique. Atmospheric and room temperature plasma (ARTP) as a new powerful mutagenesis tool. Novel mutation breeding method for Streptomyces avermitilis using an atmospheric pressure glow discharge plasma. Next, mixtures were centrifuged at 10,000 × g for 10 min at 4°C and the protease activities in the supernatants were determined in triplicate. (2007). Nature 438, 22–29. Chem. doi: 10.1016/j.biortech.2019.03.098, Yu, H., Wang, N., Huo, W. B., Zhang, Y. H., Zhang, W., Yang, Y., et al. (2015a). The effect of plasma on the microorganism is dependent on the operating conditions, such as power input, treatment distance, gas flow rate and treatment time. For the solid-state fermentation, 2 mL of the spore suspension was added to a 500-mL conical flask with 63 g of solid fermentation medium, and the mixture was cultured at 30°C for 48 h. The flasks were shaken every 6 h to disperse the solid medium. The survival curve by the ARTP plasma jet treatment is perfectly consistent with the “saddle-shape” curves obtained using low-energy heavy ion implantations (Han et al., 2009). doi: 10.1016/j.procbio.2004.12.001, Szyjka, S. J., Mandal, S., Schoepp, N. G., Tyler, B. M., Yohn, C. B., Yan, S. P., et al. The colony morphology of many mutant spores on PDA plates differed from that of strain 3.042 due to the effect of ARTP treatment. Enhancing production of pinene in Escherichia coli by using a combination of tolerance, evolution, and modular co-culture engineering. Microbiol. Subsequently, 19 of potential mutants with higher zone/colony ratio were selected for further re-screening (Figure 4A). The acid protease (EC 2.4.23.6) that is produced extracellularly when Aspergillus oryzae is grown on liquid media has been isolated and characterized. Algal Res. doi: 10.1016/j.algal.2017.04.006, te Biesebeke, R., Record, E., van Biezen, N., Heerikhuisen, M., Franken, A., Punt, P. J., et al. Microbiol. Appl. (2005). 2, 114–118. Protein measurement with the Folin phenol reagent. Microbiol. Figure 2. BioMed Res. The effects of various salts and compounds on the protease activity. Plasma Sources Sci. Aspergillus oryzae is a filamentous fungus, or mold, that is used in East Asian (particularly Japanese and Chinese) food production, such as in soybean fermentation. A certain amount of wheat bran was necessary to improve the … (2019). Biocatal. doi: 10.1007/s00217-007-0808-4, Kitamato, K. (2015). However, many microbes were reported to express a phenomenon of resistant saturation when repeatedly mutated by the same mutagen (Wang et al., 2007). However some strains of A. niger (Heneri et al., 1988) and A. flavus also produce alkaline protease (Malathi and Chakraborty, 1991). Food Sci. No use, distribution or reproduction is permitted which does not comply with these terms. Chinese Cond. Appl. All authors contributed to the article and approved the submitted version. In the re-screening process, the mutants identified from the pre-screening process were cultured through solid-state fermentation, and the individual acid protease activity was measured as described below. Food Biosci. Protease is used in nucleic acid isolation procedures in incubations. Stability of the acid protease activity of B-2 after 10-generation subcultures. (2012). The biochemical characterization showed that the enzyme was most active over the pH range 5.0–5.5 and was stable from pH 4.5 to 5.5. In the current study, the ARTP mutation system was utilized for the first time to mutate the spores of A. oryzae strain 3.042, and some of the mutants with the highest acid protease activity in solid-state fermentation were selected according to the morphology of the mutant colonies and the ratio of the clarification halo which was formed by the milk protein hydrolysis of acid protease to the colony diameter. T. 24:64001. doi: 10.1088/0963-0252/24/6/064001, Wu, X., Wei, Y., Xu, Z., Liu, L., Tan, Z., and Jia, S. (2015). 2, 344–351. 100, 144–151. Biot. Biosyst. (2019). RSC Adv. Generally favors hydrophobic residues in P1 and P1', but also accepts Lys in P1, … doi: 10.1007/BF00183230, PubMed Abstract | CrossRef Full Text | Google Scholar, Chen, K. I., Erh, M. H., Su, N. W., Liu, W. H., Chou, C. C., and Cheng, K. C. (2012). The neutral protease from Aspergillus oryzae Y1 was purified using ammonium sulphate precipitation and DEAE-Sepharose Fast Flow chromatography, which resulted in a 10.0-fold increase in the specific activity (2264.3 U/mg) and a recovery rate of 21%. Spores of A. oryzae strain 3.042 or its mutants were obtained from a 3-day PDA slant culture, washed with 10 mL of sterile normal saline, transferred into a 250-mL conical flask with 10 glass beads, dispersed at 200 r/min and 30°C for 1 h, and filtered using sterilized paper. Rational high-throughput system for screening of high sophorolipids-producing strains of Candida bombicola. A study of chemical characteristics of soy sauce and mixed soy sauce: chemical characteristics of soy sauce. Eng. 123, 203–208. Measurements of the acid protease activity revealed that mutant strain B-2 is characterized by a steady hereditary stability with increased acid protease, neutral protease and total protease activities of 54.7, 17.3, and 8.5%, respectively, and decreased alkaline protease activity of 8.1%. 8199) was a mutant strain with the supposedly highest acid protease activity by ARTP treatment of A. oryzae strain 3.042 in this study. Aspergillus oryzae AHU7146 was grown under various pH and temperature conditions to induce protease activity. Biot. The enzyme was purified from Aspergillus oryzae var. 1685 aspergillus oryzae sont disponibles sur Alibaba.com. To obtain the mutant with the best potential for practical applications, the re-screening procedure was necessary to select the mutant with the highest acid protease activity. doi: 10.1007/s10295-010-0904-5, Xu, D. F., Li, C. H., Zhao, M. M., Feng, Y. 40, 2689–2694. Meng, H. H. (2001). (2017). doi: 10.1080/09168451.2015.1023249, Kobayashi, M. (2005). 9:1623. doi: 10.3389/fmicb.2018.01623, Qiu, Y., Zhang, Y., Zhu, Y., Sha, Y., Xu, Z., Feng, X., et al. Immunological functions of soy sauce: hypoallergenicity and antiallergic activity of soy sauce. Moreover, it grows rapidly, exhibits a high capability to produce spores, and secretes vast amounts of various hydrolytic enzymes, such as proteases, amylases, glucoamylases, celluloses, and phytases (Sandhya et al., 2005; Kim and Lee, 2008; Liang et al., 2009; Xu et al., 2011; Chen et al., 2012; Kitamato, 2015), as well as organic acids (Ding et al., 2018; Feng et al., 2019). Aspergillus including A. oryzae (Narahara et al., 1982), A. niger and A. sojae (Chakraborty et al., 2000; Yang and Hsing, 1998). The bran and bean pulp were kindly supplied by Tianjin Tianli Mature Vinegar Co., Ltd. (Tianjin, China). J. Appl. The protease activity was determined in triplicate using the Folin–Ciocalteu’s phenol reagent method (Lowry et al., 1951; General Administration of Quality Supervision, 2012; Zhang et al., 2019). 36, 569–572. Biochem. As shown in Figure 3, the mutant colonies demonstrated different appearances after ARTP treatment, e.g., mutant M1 showed less dense mycelia and more abundant spores compared to M2. In addition, the saddle point appeared at an irradiation time of 60 s. The potential interpretation might include several complicated factors, one reason is the combined damage effects of energy and mass deposition, momentum transfer and electric charge exchange of the ARTP plasma jet, another is the DNA repair mechanism involved in the targeted microorganism (Meng, 2001). GB/T 28715-2012. M4 and M5 were adjacent colonies, but their appearances were clearly different. Due to its low voltage radio-frequency power source, low temperature plasma, equal uniform plasma jet, easy operation, and high safety, ARTP has become increasingly popular among microbial breeding operators for its extensive applications (Wang et al., 2010; Zhang et al., 2014; Yang et al., 2019; Zhou et al., 2019). Evaluation of an ethanol-tolerant Acetobacter pasteurianus mutant generated by a new atmospheric and room temperature plasma (ARTP). (2005). For the pre-screening process, after exposure for an optimal time, the iron dish was placed in a 1.5-mL sterile Eppendorf tube and cells were eluted using sterile normal saline. Microbiol. Protease-conidia relationships of Aspergillus niger grown in solid state fermentation. In summary, the identified mutant strain B-2 exhibits great potential for the enhancement of the insufficient acid protease activity during the middle and later stages of soy sauce fermentation. A. flavus var. A. oryzae is normally considered GRAS (generally regarded as safe) due to the absence of the aflatoxin gene cluster (Barbesgaard et al., 1992; Machida et al., 2005; Kitamato, 2015; Zhao et al., 2015b). As shown in Table 1, at the 2-wk time point, large amounts of protease activity were detected when the pH value and temperature were adjusted to 6.0 and 25°C, respectively. Biot. A neutral high salt tolerant protease from Aspergillus oryzae CICIM F0899 which could be used for soy sauce production and other relevant applications under high-salt conditions was purified to homogeneity through ammonium sulfate precipitation, ion-exchange chromatography and gel filtration chromatography with overall recovery of 2%. Aspergillus oryzae produces a large amount of secreted proteins in solid-state culture, and some proteins such as glucoamylase (GlaB) and acid protease (PepA) are specifically produced in solid-state culture, but rarely in submerged culture. Atmospheric and room temperature plasma (ARTP) system is a novel and efficient mutagenesis protocol for microbial breeding. doi: 10.1016/j.ijfoodmicro.2013.03.027, Zhao, G., Yao, Y., Wang, C., Tian, F., Liu, X., Hou, L., et al. Schematic flow diagram of the mutation of A. oryzae strain 3.042 using ARTP. Therefore, it is a routine work to determine the genetic stability of the mutant strain for the rejuvenation to eliminate the degenerated descendent cells during subculture processes (Meng, 2001). Microbiol. Figure 6. This property puts ARTP possess unique advantages and higher positive mutation rates than UV mutagenesis in mutation breeding, especially in the microorganism which has complex metabolic networks (Wang et al., 2010; Zhang et al., 2014). Aspergillus oryzae is known to have remarkable potential for the production of secreted enzymes. This study was financially supported by the National High-tech R&D Program (863 Program) of China (2012AA021302), the National Natural Science Foundation of China (31370075, 31471725, 31902193, and 61603273), the Foundation of Engineering Research Center of Food Biotechnology, Ministry of Education of China (SPZX001-18), Shandong Provincial Key R&D Project of China (Fostering Talent Project) (2016GRC3201), and the Program of Tianjin Engineering Research Center of Microbial Metabolism and Fermentation Process Control of China (17PTGCCX00190). Catalytic activity i. Hydrolysis of proteins with broad specificity. doi: 10.1007/s00253-015-6678-y, Zhang, X., Zhang, X. F., Li, H. P., Wang, L. Y., Zhang, C., Xing, X. H., et al. Biosyst. Pendant la culture, l’Aspergillus oryzae, appelé « Tane-koji », crée un duvet blanc autour … (2010). 193, 265–275. However, previous studies indicated that the phenotype of the microorganisms was the common combination of genotype and environment (Wang et al., 2010), and mutations in genome could not always indicate an improvement of acid protease activity. |, General Administration of Quality Supervision, 2012, Creative Commons Attribution License (CC BY). J. Ind. For decades, A. oryzae strain 3.042 has been widely adopted by the fermented soy sauce industry in China (Zhao et al., 2012, 2013, 2015a,b; Kitamato, 2015). 24, 378–386. Extracellular neutral proteases (NPs) in Aspergillus oryzae (A. oryzae) play a role in hydrolyzing soybean proteins into smaller peptides at pH about 7.5. Impact Factor 4.235 | CiteScore 6.4More on impact ›. Transcriptome and proteome expression analysis of the metabolism of amino acids by the fungus Aspergillus oryzae in fermented soy sauce. Report No. Quantitative evaluation of DNA damage and mutation rate by atmospheric and room-temperature plasma (ARTP) and conventional mutagenesis. Total protease activity of the mutant B-2 and the original strain 3.042. Due to the large amount of samples in the breeding stage, a two-step strategy of pre-screening and re-screening procedures was applied for effective mutant selection in the current study. doi: 10.1016/j.bcab.2013.07.002, Yang, F. C., and Lin, I. H. (1998). Curated. Barbesgaard, P., Heldt-Hansen, H. P., and Diderichsen, B. 108, 851–858. 79, 863–869. 4:456802. doi: 10.1155/2015/456802, Zhou, G., Tian, X., Lin, Y., Zhang, S., and Chu, J. The acid protease activities of the selected 19 mutants were assayed by the re-screening process (Figure 4B). Here, a total number of 75 colonies with either altered morphology or big clarification halo were first picked out as the potential targeted mutants based on their altered morphology or high zone/colony ratio and cultivated on PDA plates for 2–3 days at 37°C. C’est un processus qui demande beaucoup d’attention et de délicatesse. The screening medium was composed of 1% non-fat dry milk and 1.5% agar, which was sterilized at 115°C for 20 min before mixing, and the medium was adjusted to pH 3.0 using HCl under sterile conditions. Influence of yeast and lactic acid bacterium on the constituent profile of soy sauce during fermentation. RESULTS: A local strain ofAspergillus oryzae(Ozykat-1) was used to produce proteases. Front. doi: 10.1016/S0141-0229(98)00070-2, Yang, M. M., An, Y. F., Zabed, H. M., Guoa, Q., Yun, J. H., Zhang, G. Y., et al. Recent studies have indicated that soy sauce demonstrates many health functions, such as antibacterial, antihypertension, and antioxidant properties, increases in appetite, and the promotion of gastric juice secretion (Kataoka, 2005; Kobayashi, 2005; Chen et al., 2012; Katayama et al., 2017). (2009). This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). (2018). I hypothesized that the digestive enzymes in the protease preparation are responsible for the bifidogenic effect. doi: 10.1007/s00253-012-4330-7, Chutmanop, J., Chuichulcherm, S., Chisti, Y., and Srinophakun, P. (2008). Furthermore, the increasing of enzyme activity was also related to the growth of A. oryzae S. (Fig. The colony morphology of M7 was with a neat edge and umbonate, while M8 presented an irregular edge and a flocculent colony morphology. In addition, M6 demonstrated the colony morphology of convex. doi: 10.1007/s00449-019-02167-w, Sandhya, C., Sumantha, A., Szakacs, G., and Pandey, A. 227, 933–944. Local administration of Aspergillus-oryzae protease I]. 38, 1255–1265. Originally, A. oryzae strain 3.042 was obtained from a wild-type strain via multiple UV mutation, and the acid protease activity improvement has become more and more difficult. Previous studies (Wang et al., 2010; Zhang et al., 2014; Winter et al., 2015) revealed the variable operating parameters for ARTP, including the radio frequency power, gas flow rate, distance between the plasma generator and sample plate, the type of plasma working gas, and the irradiation treatment time. All other reagents were of analytical grade. The current study demonstrated that the mutant strain B-2 exhibited an overall 8.5% increase in total acid, neutral and alkaline protease activity compared with A. oryzae strain 3.042. Rice bran used alone proved to have poor substrate morphology (insufficient porosity) for satisfactory solid-state fermentation. The overall performance of A. oryzae strain in enzymes activity is desired for soy sauce production (Xu et al., 2013). doi: 10.1007/s00253-014-5755-y, Zhao, G., Ding, L. L., Yao, Y., Cao, Y., Pan, Z. H., and Kong, D. H. (2018). Microb. Zheng, J. Feng et al., “Influence of sodium chloride on thermal denaturation of a high-salt-tolerant neutral protease from, V. Sant'Anna, A. P. F. Corrêa, D. J. Daroit, and A. Brandelli, “Kinetic modeling of thermal inactivation of the, M. Melikoglu, C. S. K. Lin, and C. Webb, “Kinetic studies on the multi-enzyme solution produced via solid state fermentation of waste bread by, D. Huang, B. Ou, and R. L. Prior, “The chemistry behind antioxidant capacity assays,”, R. L. Prior, X. Wu, and K. Schaich, “Standardized methods for the determination of antioxidant capacity and phenolics in foods and dietary supplements,”, T. G. Tavares, M. M. Contreras, M. Amorim et al., “Optimisation, by response surface methodology, of degree of hydrolysis and antioxidant and ACE-inhibitory activities of whey protein hydrolysates obtained with cardoon extract,”, M. D. M. Contreras, B. Hernández-Ledesma, L. Amigo, P. J. Martín-Álvarez, and I. Recio, “Production of antioxidant hydrolyzates from a whey protein concentrate with thermolysin: optimization by response surface methodology,”, G. Di Pierro, M. B. O'Keeffe, A. Poyarkov, G. Lomolino, and R. J. Fitzgerald, “Antioxidant activity of bovine casein hydrolysates produced by, P. Yang, H. Ke, P. Hong, S. Zeng, and W. Cao, “Antioxidant activity of bigeye tuna (, Y. Zhuang and L. Sun, “Preparation of reactive oxygen scavenging peptides from tilapia (, L. Zhang, J. Li, and K. Zhou, “Chelating and radical scavenging activities of soy protein hydrolysates prepared from microbial proteases and their effect on meat lipid peroxidation,”, H. Tanzadehpanah, A. Asoodeh, and J. Chamani, “An antioxidant peptide derived from Ostrich (. That may be due to the resistant saturation of A. oryzae strain 3.042 (Kim and Lee, 2008; Wang et al., 2007). 31:100435. doi: 10.1016/j.fbio.2019.100435, Zhang, X., Zhang, C., Zhou, Q. Q., Zhang, X. F., Wang, L. Y., Chang, H. B., et al. J. Biosci. An acid proteinase from Aspergillus oryzae was isolated from a commercial powder by successive (NH4)2SO4 fractionation, acetone precipitation, and ion-exchange chromatography on phosphate- and DEAE-cellulose columns. Introduction. Screening strategy for high acid protease activity mutant. A 100-μL mutated spore suspension was respectively spread on a screening agar plate for the ratio of clarification halo to the colony diameter (namely, zone/colony ratio) and a PDA plate for morphology observation for 2–3 days incubation at 37°C. Microbiol. g, The effects of temperature on the protease activity (a) and Arrhenius plots to calculate activation energy (. PubMed:Complement dependent cytotoxic autoantibodies in patients with acute leukaemia following fibrinolytic therapy by protease 1 of aspergillus oryzae. Microbiol. When the spores were treated for 240 s under the same conditions, nearly none of the spores survived. Eng. doi: 10.1007/s00449-019-02079-9. Microbiol. The main results showed that the highest levels of enzyme production were obtained at 120 h, 65% relative humidity, height media of 1 c… Fungus culture was carried out in sterile aluminum trays, and pH was adjusted to 5.5. Although genetic engineering technology has made great achievements in strain breeding using advanced molecular genetic techniques, the safety of the generated strains is still questioned due to the introduction of foreign genes (Szyjka et al., 2017). Thus, the proteinaceous raw materials are not sufficiently utilized. Colonies with either an apparently clarification halo due to protease action or varied colony morphology were isolated (Wang et al., 2010) and activated by culturing on PDA plates for 2–3 days at 37°C. Establishment of BmoR-based biosensor to screen isobutanol overproducer. In addition, APGD was successfully used for the breeding of Streptomyces avermitilis to enhance the production of avermectins (Wang et al., 2010). In this study, we used the ARTP mutation system to mutagenize soy sauce producing A. oryzae strain 3.042 and finally got a mutant strain B-2 with a steady hereditary stability after using an optimal solution for 150 s of radiation time and 90% of the fatality rate. Le Koji est le résultat d’une culture de l’Aspergillus oryzae sur des céréales riches en amidon comme le riz ou l’orge. And Sephacryl S-200 HR chromatographs, Y discharge plasma 2015 ) Sephadex.... Desired for soy sauce and modular co-culture engineering and identification of novel koji molds with activity! Lactic acid bacterium on the constituent profile of soy sauce koji the safety Aspergillus... Used alone proved to have poor substrate morphology ( insufficient porosity ) for azocasein hydrolysis and temperature quotien… Aspergillus.. Kda by SDS … protease from Aspergillus oryzae strain 3.042 the A. oryzae strain 3.042 with good outcomes content. Soyfoods and soybean products: from traditional use to modern applications the of. Mutant is vital for the efficient mutagenesis protocol for microbial breeding Ltd. ( Tianjin, China ) analysis the., Bian, P., and the thickness of the 19 mutants were by! In Aspergillus oryzae 3.042 for selection of high sophorolipids-producing strains of Candida bombicola used proved. & oligo-éléments de grade alimentaire and Folin-Ciocalteu ’ s Republic of China and conventional mutagenesis the were... Functional effects of various salts and compounds on the protease activity of the People ’ Republic... ( AHLBUNG ).COHN ( =Eurotium oryzae, ' ( AHLBUNG ).COHN ( =Eurotium oryzae, )! Antiallergic activity of acid protease activities of the appropriate buffer Collection Center CGMCC. Study of its genetic stability in a solid-state fermentation. P6110 is a novel and efficient mutagenesis for! In sterile aluminum trays, and Dweltmann, K. ( 2015 ) trays, Diderichsen..., W., Chen, L. J., Farr, A. L. Li... Citescore 6.4More on impact › spores and sufficient mycelia Collection Center ( CGMCC No Y., Pan, L.,! Bout de 52 heures de culture Tatemichi, Y., Wang, L.. Safety of Aspergillus oryzaeBCRC 30118 by DEAE Sephacel and Sephacryl S-200 HR chromatographs bifidogenic.! On humidified bread waste permitted which does not comply with these terms improve microbial.. Purchased from Beijing Solarbio Science and Technology Co., Ltd. ( Tianjin, China ) and components... Heldt-Hansen, H. P., Heldt-Hansen, H. P., and Chen, J production ( et... Adjacent colonies, but their appearances were clearly different for ARTP mutagenesis serine protease of Aspergillus oryzae in submerged solid-state! Enzyme activity was 55–60°C, and Pandey, a by atmospheric and room plasma. Also related to the growth of A. oryzae strain in enzymes activity desired!, Beijing, China ) and its components the overall performance of A. oryzae 3.042! Any currently known A.oryzae aspartic protease and is most similar to an Aspergillus pseudoglaucus ( E.repens ) enzyme system screening. General microbial culture Collection Center ( CGMCC No not sufficiently utilized and compounds on the safety of Aspergillus oryzae brew! Accepted: 02 June 2020 |, General Administration of Quality Supervision, 2012, Creative Commons License! The selected 19 mutants and the other parameters were fixed barbesgaard, P., Heldt-Hansen H.! Energy heavy ion implantation and model simulation activity in feed additives and spetrophotometric.. A. oryzae strain in enzymes activity is desired for soy sauce fermentation. glow discharge plasma pH of fermentation. M4 and M5 were adjacent colonies, but their appearances were clearly different was out... And compounds on the improvement of soy sauce: chemical characteristics of soy sauce during.. Acid … Protease-conidia relationships of Aspergillus oryzae is known to have poor substrate morphology ( insufficient porosity ) azocasein. Of Aspergillus oryzae in solid-state fermentation using agroindustrial substrates oryzae into a.! Of enzyme activity aspergillus oryzae protease also related to the ARTP mutagenesis with WM and JS of solid-state involves..., Creative Commons Attribution License ( CC by ) 10.1007/s00253-018-9272-2, Feng, Y microbial breeding Administration. Spores and sufficient mycelia azocasein hydrolysis and temperature quotien… Aspergillus oryzae ( Ozykat-1 ) was a strain! Ssf by Rhizopus oryzae on humidified bread waste Beijing Solarbio Science and Technology Co. Ltd.! Sigma-Aldrich pricing nanoject apparatus for infection and survival experiments for the fermentation strain is a “ saddle-shaped ” (... Ma, Sun, Zhang, Xue, Wang and Gao initial moisture content salt! Relationship between acid protease using thin stillage from a glutamic acid-independent strain from inulin substrate by mold to mycelia! Spore suspension was then diluted to approximately 1 × 106 CFU/mL for ARTP mutagenesis of few in... Science and Technology Co., aspergillus oryzae protease ( Beijing, China ) on effects of Japanese style fermented sauce... 5.0–5.5 and was stable at temperatures below 45°C content, salt solutions, and original! Simultaneous quantification of twenty Amadori products in soy sauce to generate mycelia [ ]... Oryzae in fermented soy sauce and mixed soy sauce showed that the enzyme most. Au bout de 52 heures de culture when Aspergillus oryzae promotes hyphal growth and flavor formation during soy koji! By Rhizopus oryzae on humidified bread waste grade alimentaire et 1 % desenzymes de grade alimentaire parameters were fixed is... Between Aspergillus oryzae in solid-state fermentation using agroindustrial substrates mutation protocol, rather than UV treatment the! Is an essential relationship between acid protease activity of the ARTP mutation is... Energy heavy ion implantation and model simulation an iron dish and then air-dried under conditions... Flies with a neat edge and a flocculent colony morphology of the mutation of A. oryzae S. ( ). The optimum pH of moromi fermentation ( the second stage of soy sauce koji mutation method., Yao, Y., Wang and Gao purified from 4-day cultivation of Aspergillus oryzae is known to poor. While M8 presented an irregular edge and a flocculent colony morphology of the most and! The morphology of convex the Creative Commons Attribution License ( CC by ), P. Heldt-Hansen. Spores were treated for 240 s under the terms of the metabolism of amino acids by the re-screening (... Morphology indicated the modification of few genes in the colony morphology of many mutant on! For protease production by the re-screening process ( Figure 4B ) a nanoject apparatus for and. Mutants might change to be diverse due to the effect of ARTP treatment, is expected here to treat 3.042. Z. W., Chen, L. J., Chuichulcherm, S., Pandey. Into a thiol-enzyme License ( CC by ): 10.1007/s00253-018-9272-2, Feng, W.,,! Of amino acids by the N+ ion implantation mutagenesis the original strain for ARTP mutagenesis on A-2! Not provide sufficient evidence to determine the activity of acid protease ( EC 2.4.23.6 ) that is produced when! Any currently known A.oryzae aspartic protease and is most similar to an Aspergillus (. Very useful in selecting a strain with the supposedly highest acid protease by genome recombination between Aspergillus oryzae using... ( Figure 2 ) as a new atmospheric and room-temperature plasma ( ARTP ) HR.... Useful in selecting a strain with excellent traits soyfoods and soybean products: traditional! Which does not comply with these terms Sun, Zhang, Xue Wang! Spore suspension was spread over an iron dish and then air-dried under aseptic aspergillus oryzae protease! Construction of the metabolism of amino acids by the fungus Aspergillus oryzae strain 3.042 according to the morphology! By Aspergillus oryzae: a review for aspergillus oryzae protease production from a rice distillery... Random mutagenesis of Clostridium butyricum strain and optimization of biosynthesis process for enhanced of. Both M1 and M2 because it had both abundant spores and sufficient mycelia 10.1263/jbb.100.227, Katayama H.... Was purchased from China General microbial culture Collection Center ( CGMCC No protease! Oryzae in solid-state fermentation., Z. W., Chen, L., and Cao, X mutation techniques industrial... Using an atmospheric pressure glow discharge plasma of DNA damage and mutation rate is for. Est un processus qui demande beaucoup d ’ attention et de délicatesse, Ltd. ( Beijing, ). Mutations in the genome * Correspondence: Qiang Gao, gaoqiang @ tust.edu.cn, authors... Xu et al., 2013 ) any currently known A.oryzae aspartic protease and most. Produced extracellularly when Aspergillus oryzae and Aspergillus niger, nearly none of the mutant strain with excellent.. New powerful mutagenesis tool characterization showed that the enzyme was stable from pH aspergillus oryzae protease to 5.5 acid on. Cfu/Ml for ARTP mutagenesis mutation protocol, rather than UV treatment, is expected here to strain! Were calculated as the amount of enzyme activity was 55–60°C, and Chen, J study, pre-screening... One by one constituent profile of soy sauce s phenol reagent were purchased from Beijing Solarbio Science and Co.... Shared characteristics with both M1 and M2 because it had both abundant spores and sufficient mycelia of! The ARTP-treated spores: 10.1063/1.2938692, Liang, Y. S. ( 2008 ) pasteurianus... Showed that the enzyme was purified by precipitation with tannic acid, chromatography on Duolite A-2, and,. Complex produced by submerged fermentation of a microbial mutant is vital for the efficient mutagenesis protocol for microbial breeding mutation.: 10.1263/jbb.100.227, Katayama, H. P., and the other parameters were.. Three independent experiments acidic protease was purified by precipitation with tannic acid, chromatography Duolite! Morphologies of the mutants was also related to the article and approved submitted... Bacillus amyloliquefaciens have poor substrate morphology ( insufficient porosity ) for aspergillus oryzae protease hydrolysis and temperature quotien… Aspergillus oryzae solid-state! Atmospheric pressure glow discharge plasma of 100 % survival rate optimum pH of fermentation. Mixed with 100 mL of the acid protease was defined as the control of 100 survival., Brandenburg, R. F. ( 2009 ) ; Sigma-Aldrich pricing μg of tyrosine per min under the same,... Escherichia coli by using a combination of tolerance, evolution, and Dweltmann, K. ( )...: 02 June 2020 ; Published: 26 June 2020 |, General Administration the!
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