Fungus

Technical Data

Type of microorganism	Fungus
Target proteins	Dairy proteins, egg proteins, enzymes. See organism table for specific cases.
Wild-type or GMO	Mostly GMO, also one wild-type case. See organism table for specific cases.
Production mode (intracellular/extracellular)	Mainly extracellular production due to ease of DSP (Eastham & Leman, 2024)
C & N source	Mainly glucose and inorganic sources of N used (ammonia, urea, nitrate), but fungi can use a wide variety of C & N sources, including industrial waste streams (Lübeck & Lübeck, 2022). See organism table for specific cases.
Regulatory status in Europe	The production of food related proteins in fungi is not allowed. One case is issued as safe for consumption by EFSA, but is not allowed yet. See organism table for specific cases.
Regulatory status in other parts of the world	Several products have FDA GRAS apporval in the US. No product allowed in Canada. See organism table for specific cases.
Companies	Perfect Day Onego Bio Danisco Chr. Hansen (part of Novonesis) Novozymes (part of Novonesis)
Average yield	1-32 g/L in published reports, 5-100 g/L in industrial context (Eastham & Leman, 2024)
General temperature range	20-30°C (Zhuang et al., 2024)
General pH range	pH 5.5-8 (Zhuang et al., 2024)
Growth rate (µ)	0.12-0.35/hour (Bajić et al., 2022)
Ease of genetic modification	A lot of tools are present. Some problems with low efficiency because of their cell wall and preference of NHEJ over HR, but has recently been improved by CRISPR-Cas. (P. Yang et al., 2024)
Feedstock suitability	Wide variety of agro-industrial waste streams can be used as feedstock (Rajput et al., 2024). See organism table for specific cases.
Downstream purification processing complexity (isloation, lysis, purification)	For secreted proteins: simple centrifugation followed by purification of protein in the supernatant (mostly done by chromatography, complexity depends on the protein) (Tripathi & Shrivastava, 2019) For intracellular & periplasmic proteins: cell lysis step needed. Fungi are generally harder to lyse than bacteria and yeasts due to their very rigid cell wall (Gow et al., 2017)
Advantages	Eukaryotic PTMs, good protein folding, high secretion levels, high yields (Knychala et al., 2024)
Challenges (Key limitations, risk factors)	Secrete proteases, cell lysis during fermentation can release intracellular proteases, glycosylation patterns are diferent form mammals (Knychala et al., 2024)
Publications/references	Eastham, J. L., & Leman, A. R. (2024). Precision fermentation for food proteins: ingredient innovations, bioprocess considerations, and outlook — a mini-review. Current Opinion in Food Science, 58, 101194. https://doi.org/10.1016/j.cofs.2024.101194 Lübeck, M., & Lübeck, P. S. (2022). Fungal Cell Factories for Efficient and Sustainable Production of Proteins and Peptides. Microorganisms, 10(4), 753. https://doi.org/10.3390/microorganisms10040753 Knychala, M. M., Boing, L. A., Ienczak, J. L., Trichez, D., & Stambuk, B. U. (2024). Precision Fermentation as an Alternative to Animal Protein, a Review. Fermentation, 10(6), 315. https://doi.org/10.3390/fermentation10060315 Zhuang, Z., Wan, G., Lu, X., Xie, L., Yu, T., & Tang, H. (2024). Metabolic engineering for single-cell protein production from renewable feedstocks and its applications. Advanced Biotechnology, 2(4). https://doi.org/10.1007/s44307-024-00042-8 Rajput, S. D., Pandey, N., & Sahu, K. (2024). A comprehensive report on valorization of waste to single cell protein: strategies, challenges, and future prospects. Environmental Science and Pollution Research, 31(18), 26378–26414. https://doi.org/10.1007/s11356-024-33004-7 Bajić, B., Vučurović, D., Vasić, Đ., Jevtić-Mučibabić, R., & Dodić, S. (2022). Biotechnological Production of Sustainable Microbial Proteins from Agro-Industrial Residues and By-Products. Foods, 12(1), 107. https://doi.org/10.3390/foods12010107 Yang, P., Condrich, A., Lu, L., Scranton, S., Hebner, C., Sheykhhasan, M., & Ali, M. A. (2024). Genetic Engineering in Bacteria, Fungi, and Oomycetes, Taking Advantage of CRISPR. DNA, 4(4), 427–454. https://doi.org/10.3390/dna4040030 Tripathi, N. K., & Shrivastava, A. (2019). Recent Developments in Bioprocessing of Recombinant Proteins: Expression Hosts and Process Development. Frontiers in Bioengineering and Biotechnology, 7. https://doi.org/10.3389/fbioe.2019.00420 Gow, N. a. R., Latge, J., & Munro, C. A. (2017). The Fungal Cell Wall: Structure, Biosynthesis, and Function. Microbiology Spectrum, 5(3). https://doi.org/10.1128/microbiolspec.funk-0035-2016

Trichoderma reesei

Aspergillus niger