title: Consolidated conversion of protein waste into biofuels and ammonia using Bacillus subtilis.
abstract: The non-recyclable use of nitrogen fertilizers in microbial production of fuels and chemicals remains environmentally detrimental. Conversion of protein wastes into biofuels and ammonia by engineering nitrogen flux in Escherichia coli has been demonstrated as a method to reclaim reduced-nitrogen and curb its environmental deposition. However, protein biomass requires a proteolysis process before it can be taken up and converted by any microbe. Here, we metabolically engineered Bacillus subtilis to hydrolyze polypeptides through its secreted proteases and to convert amino acids into advanced biofuels and ammonia fertilizer. Redirection of B. subtilis metabolism for amino-acid conversion required inactivation of the branched-chain amino-acid (BCAA) global regulator CodY. Additionally, the lipoamide acyltransferase (bkdB) was deleted to prevent conversion of branched-chain 2-keto acids into their acyl-CoA derivatives. With these deletions and heterologous expression of a keto-acid decarboxylase and an alcohol dehydrogenase, the final strain produced biofuels and ammonia from an amino-acid media with 18.9% and 46.6% of the maximum theoretical yield. The process was also demonstrated on several waste proteins. The results demonstrate the feasibility of direct microbial conversion of polypeptides into sustainable products.
authors.affiliation: Department of Chemical and Biomolecular Engineering, University of California, 7523 Boelter Hall, 420 Westwood Plaza, Los Angeles, CA 90095, USA; Department of Environmental Engineering, College of Engineering, Ajou University, Suwon, Gyeonggi-do, South Korea.
authors.forename: Kwon-Young
authors.initials: KY
authors.lastname: Choi
authors.affiliation: Department of Chemical and Biomolecular Engineering, University of California, 7523 Boelter Hall, 420 Westwood Plaza, Los Angeles, CA 90095, USA.
authors.forename: David G
authors.initials: DG
authors.lastname: Wernick
authors.affiliation: Department of Chemical and Biomolecular Engineering, University of California, 7523 Boelter Hall, 420 Westwood Plaza, Los Angeles, CA 90095, USA.
authors.forename: Christine A
authors.initials: CA
authors.lastname: Tat
authors.affiliation: Department of Chemical and Biomolecular Engineering, University of California, 7523 Boelter Hall, 420 Westwood Plaza, Los Angeles, CA 90095, USA; Department of Chemistry and Biochemistry, University of California, 607 Charles E. Young Drive East, Los Angeles, CA 90095, USA; Institute for Genomics and Proteomics, University of California, 201 Boyer Hall, 611 Charles E. Young Drive East, Los Angeles, CA 90095, USA; The Molecular Biology Institute, University of California, Paul D. Boyer Hall Box 951570, 611 Charles E. Young Drive East, Los Angeles, CA 90095, USA. Electronic address: liaoj@seas.ucla.edu.
authors.forename: James C
authors.initials: JC
authors.lastname: Liao
mesh_terms: D000641:Ammonia / Q000378:metabolism*; D001412:Bacillus subtilis / Q000235:genetics / Q000378:metabolism; D056804:Biofuels; D060847:Metabolic Engineering / Q000379:methods; D011506:Proteins / Q000378:metabolism*; D014873:Water Pollutants / Q000378:metabolism*; D014876:Water Pollution; D018508:Water Purification / Q000379:methods*
chemical_list: D056804:Biofuels; D011506:Proteins; D014873:Water Pollutants; D000641:Ammonia
keywords: Ammonia; BCKDC; Bacillus subtilis; CodY; Higher alcohols; Protein biomass; Protein conversion