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BIONÆR-Bionæringsprogram

Thermophilic break-down of keratin-laden biomass waste

Alternative title: Termofil bakteriell nedbrytning av keratin-holdige biologiske avfallsprodukter

Awarded: NOK 4.3 mill.

Keratin is a fibrous and recalcitrant structural protein and is the third most abundant polymer in nature after cellulose and chitin. Animals and birds have developed a diversity of keratins used as structural parts of their outer protection which make up major components of feathers, hair, horns, hooves, cloves, nails etc. Their recalcitrant nature is due to a high degree of cross-linking chemical bonds, making them relatively resistant to biological, physical, and chemical degradation. Keratin-laden tissues represent a significant challenge for the animal rendering industry. For example, feathers consist of more than 90% keratin and represent a huge waste product of the poultry industry, where most ends up in landfills or is being burned. Similar challenges exist for other keratin-containing biomass waste. Today, most feather waste is discarded but some is ineffectively rendered for animal feed or fertilizer. This project addresses the application of selected anaerobic heat-living bacteria which can be optimized for keratin-laden waste material degradation and the understanding of the enzyme activities within the bacterial species responsible for this degradation. In the first phase of this project, we examined a range of bacteria belonging to the genus Fervidobacterium for their feather-degrading capacity. Seven species were identified as being particularly active. We have elucidated the complete genome sequences of six species/strains by combining Oxford Nanopore Technology and Illumina Technology. These genomes have been analyzed and annotated to find potential genes related to the keratin degradation process, and compared. For three of these strains, we have completed comparative transcriptional analyses (transcriptomics), cultivating them with feather and comparing with cultures with glucose as the sole carbon and energy source, to identify genes that are expressed more strongly under keratinolytic growth. A number of overexpressed peptidase- and protease-encoding genes have been identified in these experiments in the most active strains. Similarly, proteomics analyses of cell-bound proteins and proteins secreted to the extracellular environment have identified a number of proteins secreted by the cells under keratinolytic growth conditions. By using these approaches, approximately 20 genes encoding proteases and peptidases were found to be more strongly expressed under keratinolytic conditions in the most efficient strains, indicating that these genes/enzymes play important roles in the feather-degrading process. Furthermore, by combining all the upregulated proteins found in the proteomics experiments with those overexpressed genes from the transcriptomics ones, we created a gene network for each of the strains with more than 100 overexpressed genes under keratinolytic growth conditions. These genes are likely biologically connected, and further analyses may lead to a better understanding of the keratin break-down process. This work has been done in collaboration with the French ThermoK partner. In collaboration with partners from South-Africa and UK, several of the proteases mentioned above have been produced by heterologous gene expression in Escherichia coli, purified and characterized biochemically and structurally. This project will lead to improved understanding and control of the overall keratin-degrading process and improve the use of these microorganisms and their enzymes in a cost effective and controllable manner. ThermoK brings together a multidisciplinary team of academics to develop this field and includes up-scaling of the keratin-degrading processes to larger bioreactors. The project will contribute to the designing of more sustainable and resilient food systems and contribute to the vision of a circular economy by using waste products and converting them to other valuable commercial products including proteins, peptides, amino acids, feed and fertilizers.

Keratin is a fibrous and recalcitrant structural protein and is the third most abundant polymer in nature after cellulose and chitin. A wide spectrum of animals have developed a diversity of keratins used as structural parts of their outer protection which make up major component of feathers, hair, horns, hooves, cloves, nails etc. Their recalcitrant nature is due to properties such as a high degree of cross-linking by disulphide bonds, hydrogen bonds and hydrophobic interactions. Keratin-laden tissues represents a significant challenge for the animal rendering industry. For example, feathers consist of more than 90% keratin and represent a huge waste product of the poultry industry, where most ends up in landfills or is being burned. Similar challenges exist for other keratin-containing biomass waste. Today, most feather waste is discarded or ineffectively rendered for animal feed or fertilizer. This project will address the application of selected anaerobic thermophilic bacteria which can be optimized for keratin-laden waste material degradation as well as the understanding of the enzyme activities within the bacterial species responsible for this degradation. This will lead to improved control and understanding of the overall keratin-degrading process and its improvement and efficiency by using organisms expressing the required activities or using novel enzyme cascades of thermophilic keratin degrading enzymes in vitro optimised for keratin breakdown in a cost effective and controllable manner. ThermoK will bring together a multidisciplinary team of academics in order to take the results obtained to a transfer level 5 and above, including upscaling of the process. The project will contribute to the designing of more sustainable and resilient food systems and contribute to the vision of a circular economy by using waste products and converting them to other valuable commercial products including peptides, amino acids, fish feed and agricultural fertilizers.

Funding scheme:

BIONÆR-Bionæringsprogram