Interspecific protoplast fusion of atmospheric and room-temperature plasma mutants of Aspergillus generates an L-asparaginase hyper-producing hybrid with techno-economic benefits

References

• Vala, A. K.; Sachaniya, B.; Dudhagara, D.; Panseriya, H. Z.; Gosai, H.; Rawal, R.; Dave, B. P. Characterization of L-Asparaginase from Marine-Derived Aspergillus niger AKV-MKBU, Its Antiproliferative Activity and Bench Scale Production Using Industrial Waste. Int. J. Biol. Macromol. 2018, 108, 41–46. DOI: 10.1016/j.ijbiomac.2017.11.114.
   PubMed Web of Science ®Google Scholar
• Shakambari, G.; Kumar, R. S.; Ashokkumar, B.; Varalakshmi, P. Agro Waste Utilization for Cost-Effective Production of L‑Asparaginase by Pseudomonas plecoglossicida RS1 with Anticancer and Acrylamide Mitigation Potential. ACS Omega 2017, 2, 8108–8117. DOI: 10.1021/acsomega.7b01429.
   PubMed Web of Science ®Google Scholar
• Mihooliya, K. N.; Nandal, J.; Kumari, A.; Nanda, S.; Verma, H.; Sahoo, D. K. Studies on Efficient Production of a Novel L‑Asparaginase by a Newly Isolated Pseudomonas resinovorans IGS‑131 and Its Heterologous Expression in Escherichia coli. 3 Biotech 2020, 10, 148–158. DOI: 10.1007/s13205-020-2135-4.
   PubMedGoogle Scholar
• Al‑Hazmi, N. E.; Naguib, D. M. Plant Asparaginase versus Microbial Asparaginase as Anticancer Agent. Environ. Sci. Pollut. Res. Int. 2022, 29, 27283–27293. DOI: 10.1007/s11356-021-17925-1.
   PubMed Web of Science ®Google Scholar
• Cachumba, J. J. M.; Antunes, F. A. F.; Peres, G. F. D.; Brumano, L. P.; Santos, J. C. D.; Da Silva, S. S. Current Applications and Different Approaches for Microbial L-Asparaginase Production. Braz. J. Microbiol. 2016, 47, 77–85. DOI: 10.1016/j.bjm.2016.10.004.
   PubMed Web of Science ®Google Scholar
• Ekpenyong, M. G.; Asitok, A. D.; Antigha, R. E.; Ogarekpe, N. M.; Ekong, U. S.; Asuquo, M. I.; Essien, J. P.; Antai, S. P. Bioprocess Optimization of Nutritional Parameters for Enhanced anti-Leukemic L-Asparaginase Production by Aspergillus candidus UCCM 00117: A Sequential Statistical Approach. Int. J. Pept. Res. Ther. 2021, 27, 1501–1527. DOI: 10.1007/s10989-021-10188-x.
   PubMed Web of Science ®Google Scholar
• Chan, W. K.; Horvath, T. D.; Tan, L.; Link, T.; Harutyunyan, K. G.; Pontikos, M. A.; Anishkin, A.; Du, D.; Martin, L. A.; Yin, E.; et al. Glutaminase Activity of L-Asparaginase Contributes to Durable Preclinical Activity against Acute Lymphoblastic Leukemia. Mol. Cancer Ther. 2019, 18, 1587–1592. 10.1158/1535-7163.
   PubMed Web of Science ®Google Scholar
• Lopes, A.; Nascimento, L.; Ribeiro, A.; Tairum, C., Jr; Breyer, C.; de Oliveira, M.; Monteiro, G.; de Souza-Motta, C.; Magalhães, P.; Avendaño, J.; et al. Therapeutic L-Asparaginase: Upstream, Downstream and beyond. Crit. Rev. Biotechnol. 2017, 37, 82–99. DOI: 10.3109/07388551.2015.1120705.
   PubMed Web of Science ®Google Scholar
• Sadh, P. K.; Kumar, S.; Chawla, P.; Duhan, J. S. Fermentation: A Boon for Production of Bioactive Compounds by Processing of Food Industries Wastes (by-Products). Molecules 2018, 23, e2560. DOI: 10.3390/molecules23102560.
   PubMed Web of Science ®Google Scholar
• Pinu, F. R.; Granucci, N.; Daniell, J.; Han, T.-L.; Carneiro, S.; Rocha, I.; Nielsen, J.; Villas-Boas, S. G. Metabolite Secretion in Microorganisms: The Theory of Metabolic Overflow Put to the Test. Metabolomics 2018, 14, 43–58. DOI: 10.1007/s11306-018-1339-7.
   PubMed Web of Science ®Google Scholar
• Zou, R.-S.; Li, S.; Zhang, L.-L.; Zhang, C.; Han, Y.-J.; Gao, G.; Sun, X.; Gong, X. Mutagenesis of Rhodobacter sphaeroides Using Atmospheric and Room Temperature Plasma Treatment for Efficient Production of Coenzyme Q10. J. Biosci. Bioeng. 2019, 127, 698–702. DOI: 10.1016/j.jbiosc.2018.12.005.
   PubMed Web of Science ®Google Scholar
• Zhang, X.; Wang, Z.; Wang, Y.; Li, X.; Zhu, M.; Zhang, H.; Xu, M.; Yang, T.; Rao, Z. Heterologous Expression and Rational Design of L-Asparaginase from Rhizomucor miehei to Improve Thermostability. Biology 2021, 10, 1346. DOI: 10.3390/biology10121346.
   PubMedGoogle Scholar
• Katayama, T.; Maruyama, J. CRISPR/Cpf1-Mediated Mutagenesis and Gene Deletion in Industrial Filamentous Fungi Aspergillus oryzae and Aspergillus Sojae. J. Biosci. Bioeng. 2022, 133, 353–361. DOI: 10.1016/j.jbiosc.2021.12.017.
   PubMed Web of Science ®Google Scholar
• Asitok, A. D.; Ekpenyong, M. G.; Takon, I. A.; Antai, S. P.; Ogarekpe, N. M.; Antigha, R. E.; Edet, P. E.; Ben, U. C.; Akpan, A. E.; Antai, A. S.; Essien, J. P. Overproduction of a Thermo-Stable Halo-Alkaline Protease on Agro-Waste-Based Optimized Medium through Alternate Combinatorial Random Mutagenesis of Stenotrophomonas acidaminiphila. Biotechnol. Rep. 2022, 35, e00746. DOI: 10.1016/j.btre.2022.e00746.
   Google Scholar
• Zhang, X.; Zhang, X.-F.; Li, H.-P.; Wang, L.-Y.; Zhang, C.; Xing, X.-H.; Bao, C.-Y. Atmospheric and Room Temperature Plasma (ARTP) as a New Powerful Mutagenesis Tool. Appl. Microbiol. Biotechnol. 2014, 98, 5387–5396. DOI: 10.1007/s00253-014-5755-y.
   PubMed Web of Science ®Google Scholar
• Strom, N. B.; Bushley, K. E. Two Genomes Are Better than One: History, Genetics and Biotechnological Applications of Fungal Heterokaryons. Fungal Biol. Biotechnol. 2016, 3, 4–17. 10.1186/s40694-016-0022-x.
   PubMedGoogle Scholar
• Klinsupa, W.; Phansiri, S.; Thongpradis, P.; Yongsmith, B.; Pothiratana, C. Enhancement of Yellow Pigment Production by Intraspecific Protoplast Fusion of Monascus spp. Yellow Mutant (Ade(-)) and White Mutant (Prototroph). J. Biotechnol. 2016, 217, 62–71. DOI: 10.1016/j.jbiotec.2015.11.002.
   PubMed Web of Science ®Google Scholar
• Zhu, F. M.; Du, B.; Gao, H. S.; Liu, C. J.; Li, J. Purification and Characterization of an Intracellular β-Glucosidase from the Protoplast Fusant of Aspergillus oryzae and Aspergillus niger. Appl. Biochem. Microbiol. 2010, 46, 626–632. DOI: 10.1134/S0003683810060116.
   Web of Science ®Google Scholar
• El-Gendy, M. M. A. A.; Al-Zahrani, S. H. M.; El-Bondkly, A. M. A. Construction of Potent Recombinant Strain through Intergeneric Protoplast Fusion in Endophytic Fungi for Anticancerous Enzymes Production Using Rice Straw. Appl. Biochem. Biotechnol. 2017, 183, 30–50. DOI: 10.1007/s12010-017-2429-0.
   PubMed Web of Science ®Google Scholar
• Shu, L.; Si, X.; Yang, X.; Ma, W.; Sun, J.; Zhang, J.; Xue, X.; Wang, D.; Gao, Q. Enhancement of Acid Protease Activity of Aspergillus oryzae Using Atmospheric and Room Temperature Plasma. Front. Microbiol. 2020, 11, 1418. DOI: 10.3389/fmicb.2020.01418.
   PubMed Web of Science ®Google Scholar
• Imada, A.; Igarasi, S.; Nakahama, K.; Isono, M. Asparaginase and Glutaminase Activities of Microorganisms. J. Gen. Microbiol. 1973, 76, 85–99. DOI: 10.1099/00221287-76-1-85.
   PubMedGoogle Scholar
• Holliday, R. A New Method for the Identification of Biochemical Mutants of Microorganisms. Nature 1956, 178, 987–987. DOI: 10.1038/178987a0.
   Web of Science ®Google Scholar
• Mormann, S.; Lömker, A.; Rückert, C.; Gaigalat, L.; Tauch, A.; Pühler, A.; Kalinowski, J. Random Mutagenesis in Corynebacterium glutamicum ATCC 13032 Using an IS6100-Based Transposon Vector Identified the Last Unknown Gene in the Histidine Biosynthesis Pathway. BMC Genomics 2006, 7, 205–224. DOI: 10.1186/1471-2164-7-205.
   PubMed Web of Science ®Google Scholar
• Sugimoto, K.; Oda, S. Efficient Production of Fungal Spores by the Combination of Reduction of Nitrogen Source Content and Embedding of Hydrophobic Polymer in an Agar Plate. J. Biosci. Bioeng. 2021, 131, 390–395. DOI: 10.1016/j.jbiosc.2020.12.007.
   PubMed Web of Science ®Google Scholar
• Cachumba, J. J. M.; Hilares, R. T.; Brumano, L. P.; Marcelino, P. R. F.; Antunes, F. A. F.; Santos, J. C.; da Silva, S. S. Extracellular L-Asparaginase Production in Solid State Fermentation by Using Sugarcane Bagasse as Support Material. Prep. Biochem. Biotechnol. 2019, 49, 328–333. DOI: 10.1080/10826068.2019.1566152.
   PubMed Web of Science ®Google Scholar
• Bradford, M. M. A Rapid and Sensitive Method for Quantitation of Microgram Quantities of Protein Utilizing the Principle of Protein-Dye Binding. Anal. Biochem. 1976, 72, 248–254. DOI: 10.1016/0003-2697(76)90527-3.
   PubMed Web of Science ®Google Scholar
• Rodrigues, L.; Moldes, A.; Teixeira, J.; Oliveira, R. Kinetic Study of Fermentative Biosurfactant Production by Lactobacillus Strains. Biochem. Eng. J. 2006, 28, 109–116. DOI: 10.1016/j.bej.2005.06.001.
   Web of Science ®Google Scholar
• Miller, G. L. Use of Dinitrosalicylic Acid Reagent for Determination of Reducing Sugar. Anal. Chem. 1959, 31, 426–428. DOI: 10.1021/ac60147a030.
   Web of Science ®Google Scholar
• Ekpenyong, M.; Asitok, A.; Antai, S.; Ekpo, B.; Antigha, R.; Ogarekpe, N.; Antai, A.; Ogbuagu, U.; Ayara, N. Kinetic Modeling and Quasi-Economic Analysis of Fermentative Glycolipopeptide Biosurfactant Production in a Medium co-Optimized by Statistical and Neural Network Approaches. Prep. Biochem. Biotechnol. 2021, 51, 450–466. DOI: 10.1080/10826068.2020.1830414.
   PubMed Web of Science ®Google Scholar
• Yang, Y.; Sha, M. A Beginner’s Guide to Bioprocess Modes: Batch, Fed-Batch, and Continuous Fermentation. Eppendorff Appl. Note 2022, 408, 1–16.
   Google Scholar
• Nascimento, C. O.; Costa, R. M.; Soares, P. B.; Herculano, P. A.; Porto, P. N.; Thiago, P. S.; Nascimento, P.; Lima, T. P.; Bezerra, C. A.; Teixeira, R. P.; et al. Purification of a Lectin from Cratylia mollis Crude Extract Seed by a Single Step PEG/Phosphate Aqueous Two-Phase System. Prep. Biochem. Biotechnol. 2020, 50, 655–663. DOI: 10.1080/10826068.2020.1725771.
   PubMed Web of Science ®Google Scholar
• Laemmli, U. K. Cleavage of Structural Protein during the Assembly of the Head 385 of Bacteriophage T4. Nature 1970, 227, 680–685. DOI: 10.1038/227680a0.
   PubMed Web of Science ®Google Scholar
• Asitok, A.; Ekpenyong, M.; Ogarekpe, N.; Antigha, R.; Takon, I.; Rao, A.; Iheanacho, A.; Antai, S. Intracellular-to-Extracellular Localization Switch of Acidic Lipase in Enterobacter cloacae: evaluation of Production Kinetics and Enantioselective Esterification Potential for Pharmaceutical Applications. Prep. Biochem. Biotechnol. 2022, 1–15. DOI: 10.1080/10826068.2022.2114010.
   PubMed Web of Science ®Google Scholar
• Ekpenyong, M. G.; Asitok, A. D.; Antai, S. P.; Ekpo, B. O.; Antigha, R. E.; Ogarekpe, N. M. Statistical and Artificial Neural Network Approaches to Modeling and Optimization of Fermentation Conditions for Production of a Surface/Bioactive Glyco-Lipo-Peptide. Int. J. Pept. Res. Ther. 2021, 27, 475–495. DOI: 10.1007/s10989-020-10094-8.
   PubMed Web of Science ®Google Scholar
• Prior, L.; Wu, X.; Schaich, K. Standardized Methods for the Determination of Antioxidant Capacity and Phenolics in Foods and Dietary Supplements. J. Agric. Food Chem. 2005, 53, 4290–4302. DOI: 10.1021/jf0502698.
   PubMed Web of Science ®Google Scholar
• Ottenheim, C.; Nawrath, M.; Wu, J. C. Microbial Mutagenesis by Atmospheric and Room-Temperature Plasma: The Latest Development. Bioresour. Bioprocess 2018, 5, 12. DOI: 10.1186/s40643-018-0200-1.
   Web of Science ®Google Scholar
• Rao, S. S.; Mamadou, L.; McConnell, M.; Polisetty, R.; Kwanyuen, P.; Hildebrand, D. Non-Antibiotic Selection Systems for Soybean Somatic Embryos: The Lysine Analog Aminoethyl-Cysteine as a Selection Agent. BMC Biotechnol. 2009, 9, 94. DOI: 10.1186/1472-6750-9-94.
   PubMed Web of Science ®Google Scholar
• Krause, F. S.; Blombach, B.; Eikmanns, B. J. Metabolic Engineering of Corynebacterium glutamicum for 2-Ketoisovalerate Production. Appl. Environ. Microbiol. 2010, 76, 8053–8061. DOI: 10.1128/AEM.01710-10.
   PubMed Web of Science ®Google Scholar
• Ushijima, S.; Nakadai, T. Breeding by Protoplast Fusion of Koji Mold. Aspergillus Sojae. Agric. Biol. Chem. 1987, 51, 1051–1057. DOI: 10.1080/00021369.1987.10868173.
   Web of Science ®Google Scholar
• da Cunha, M. C.; Aguilar, J. G. S.; Lindo, S. M. D. R. O.; de Castro, R. J. S.; Sato, H. H. L-Asparaginase from Aspergillus oryzae spp.: Effects of Production Process and Biochemical Parameters. Prep. Biochem. Biotechnol. 2022, 52, 253–263. 10/1080/10826068.2021.1931881.
   PubMedGoogle Scholar
• Santos, J. H. P. M.; Flores-Santos, J. C.; Meneguetti, G. P.; Rangel-Yagui, C. O.; Coutinho, J. A. P.; Vitolo, M.; Ventura, S. P. M.; Pessoa, A. Jr. In Situ Purification of Periplasmatic L-Asparaginase by Aqueous Two Phase Systems with Ionic Liquids (ILs) as Adjuvants. J. Chem. Technol. Biotechnol. 2018, 93, 1871–1880. DOI: 10.1002/jctb.5455.
   Web of Science ®Google Scholar
• Cardoso, S. L.; de Freitas, M. M.; de Souza, P. M.; Homem-de-Mello, M.; Silveira, D.; Fonseca-Bazzo, Y. M.; Filho, E. X.; Junior, A. P.; Magalhães, P. O. Optimization of Aqueous Two-Phase Micellar System for Partial Purification of L-Asparaginase from Penicillium sp. grown in Wheat Bran as Agro-Industrial Residue. Braz. J. Microbiol. 2020, 51, 979–988. DOI: 10.1007/s42770-020-00269-2.
   PubMed Web of Science ®Google Scholar
• Dias, F. F. G.; Ruiz, A. L. T. G.; Torre, A. D.; Sato, H. H. Purification, Characterization and Antiproliferative Activity of L-Asparaginase from Aspergillus oryzae CCT 3940 with No Glutaminase Activity. Asian Pac. J. Trop. Biomed. 2016, 6, 785–794. DOI: 10.1016/j.apjtb.2016.07.007.
   Web of Science ®Google Scholar
• Prakash, P.; Singh, H. R.; Jha, S. K. Production, Purification and Kinetic Characterization of Glutaminase Free anti-Leukemic L-Asparaginase with Low Endotoxin Level from Novel Soil Isolate. Prep. Biochem. Biotechnol. 2020, 50, 260–271. DOI: 10.1080/10826068.2019.1692221.
   PubMed Web of Science ®Google Scholar
• El-Gendy, M. M. A. A.; Awad, M. F.; El-Shenawy, F. S.; El-Bondkly, A. M. A. Production, Purification, Characterization, Antioxidant and Antiproliferative Activities of Extracellular L-Asparaginase Produced by Fusarium equiseti AHMF4. Saudi J. Biol. Sci. 2021, 28, 2540–2548. DOI: 10.1016/j.sjbs.2021.01.058.
   PubMed Web of Science ®Google Scholar
• Sangster, J. Octonol-Water Partition Coefficients of Simple Organic Compounds. J. Phys. Chem. Ref. Data 1989, 18, 1111–1229. DOI: 10.1063/1.555833.
   Web of Science ®Google Scholar
• El-Naggar, N. E.; Deraz, S. F.; El-Ewasy, S. M.; Suddek, G. M. Purification, Characterization and Immunogenicity Assessment of Glutaminase Free L-Asparaginase from Streptomyces Brollosae NEAE-115. BMC Pharmacol. Toxicol. 2018, 19, 51–65. DOI: 10.1186/s40360-018-0242-1.
   PubMed Web of Science ®Google Scholar
• Saeed, H.; Elsawy, E.; Shalaby, M.; Abdel-Fattah, M.; Hemida, A.; Eldoksh, A.; Ataya, F. S.; Nematalla, H.; Elkewedi, M.; Labrou, N. N.; El-Nikhely, N. L-Asparaginase from Dickeya chrysanthemi: Expression, Purification and Cytotoxicity Assessment. Prep. Biochem. Biotechnol. 2022, 52, 668–680. DOI: 10.1080/10826068.2021.1983831.
   PubMed Web of Science ®Google Scholar
• Li, X.; Zhang, X.; Xu, S.; Zhang, H.; Xu, M.; Yang, T.; Wang, L.; Qian, H.; Zhang, H.; Fang, H.; et al. Simultaneous Cell Disruption and Semi-Quantitative Activity Assays for High-Throughput Screening of Thermostable L-Asparaginases. Sci. Rep. 2018, 8, 7915. DOI: 10.1038/s41598-018-26241-7.
   PubMed Web of Science ®Google Scholar
• El-Naggar, N. E.; Deraz, S. F.; Soliman, H. M.; El-Deeb, N. M.; El-Ewasy, S. M. Purification, Characterization, Cytotoxicity and Anticancer Activities of L-Asparaginase, Anti-Colon Cancer Protein, from the Newly Isolated Alkaliphilic Streptomyces fradiae NEAE-82. Sci. Rep. 2016, 6, 32926. DOI: 10.1038/srep32926.
   PubMed Web of Science ®Google Scholar
• Kafkewitz, D.; Goodman, D. L-Asparaginase Production by the Rumen Anaerobe Vibrio succinogenes. Appl. Microbiol. 1974, 24, 206–209. DOI: 10.1128/am.27.1.206-209.
   Google Scholar
• Mahajan, R. V.; Saran, S.; Kameswaran, K.; Kumar, V.; Saxena, R. K. Efficient Production of L-Asparaginase from Bacillus licheniformis with Low-Glutaminase Activity: optimization, Scale up and Acrylamide Degradation Studies. Bioresour. Technol. 2012, 125, 11–16. DOI: 10.1016/j.biortech.2012.08.086.
   PubMed Web of Science ®Google Scholar
• Hassan, S. W. M.; Farag, A. M.; Beltagy, E. A. Purification, Characterization and Anticancer Activity of L-Asparaginase Produced by Marine Aspergillus terreus. J. Pure Appl. Microbiol. 2018, 12, 1845–1854. DOI: 10.22207/JPAM.12.4.19.
   Web of Science ®Google Scholar
• Jia, R.; Wan, X.; Geng, X.; Xue, D.; Xie, Z.; Chen, C. Microbial L-Asparaginase for Application in Acrylamide Mitigation from Food: current Research Status and Future Perspectives. Microorganisms 2021, 9, 1659. DOI: 10.3390/microorganisms9081659.
   PubMed Web of Science ®Google Scholar