Ethanol is a more reduced substrate than sugars. Here, 13C‐metabolic flux analysis (MFA) revealed that ethanol catabolism could supply sufficient acetyl‐CoA and reducing equivalent for PPD biosynthesis. Then, we described modular engineering strategy to optimize a multigene pathway for protopanaxadiol (PPD) production from ethanol in Saccharomyces cerevisiae. PPD biosynthesis was divided into four modules: mevalonate (MVA) pathway module, triterpene biosynthesis module, sterol biosynthesis module, and acetyl‐CoA formation module. Combinatorially overexpressing every gene in MVA pathway and optimizing metabolic balance in triterpene biosynthesis module led to significantly enhanced PPD production (42.34 mg/L/OD600). In sterol biosynthesis module, fine‐tuning lanosterol synthase gene (ERG7) expression using TetR–TetO gene regulation system enabled further production improvement (51.26 mg/L/OD600). Furthermore, increasing cytoplasmic acetyl‐CoA supply by overexpressing a Salmonella ACS (acetyl‐CoA synthetase gene) mutant ACSseL641P improved PPD production to 66.55 mg/L/OD600. In 5 L bioreactor, PPD production of the best‐performing strain WLT‐MVA5 reached 8.09 g/L, which has been the highest titer of plant triterpene produced in yeast. © 2018 American Institute of Chemical Engineers AIChE J, 65: 866–874, 2019

A multifunctional coating with antifogging, self-cleaning, and antimicrobial properties has been prepared based on a mussel-inspired chimeric protein MP-KE, which is the first example that these proteins were successfully applied to fabricate antifogging surfaces. The coating exhibits super hydrophilic properties, as indicated by contact angles less than 5° and high light transmittance similar to bare glass substrates about 90%. The zwitterionic peptides of MP-KE empower water molecules to expand into thin hydrated films rapidly, providing the protein coating with diverse surface functions. Moreover, the coatings have excellent stability and a convenient preparation process because of the mussel adhesive motif of MP-KE which makes the coating anchor onto the surface strongly. As a protein material, this multifunctional coating possesses remarkable biocompatibility and has a potential application prospects in the biomedical and pharmaceutical fields.

A mussel-inspired chimeric protein as a multifunctional coating was engineered rationally and applied in antifouling. 3,4-Dihydroxyphenyl-alanine (Dopa) and zwitterionic peptides inspired from molecular chaperones were incorporated into the designed protein. The multifunctional protein coating can anchor onto various substrates surfaces readily and exhibits superior antifouling properties.

Ascomycin (FK520), a macrocyclic polyketide natural antibiotic, displays high anti-fungal and immunosuppressive activity. In this study, the LysR family transcriptional regulator FkbR1 was characterized, and its role in ascomycin biosynthesis was explored by gene deletion, complementation, and overexpression. Inactivation of fkbR1 led to 67.5% reduction of ascomycin production, which was restored by complementation of fkbR1. Overexpression of fkbR1 resulted in a 33.5% increase in ascomycin production compared with the parent strain FS35. These findings indicated that FkbR1 was a positive regulator for ascomycin production. Quantitative RT-PCR analysis revealed that the expressions of fkbE, fkbF, fkbS, and fkbU were downregulated in the fkbR1 deletion strain and upregulated in the fkbR1 overexpression strain. Electrophoretic mobility shift assays (EMSAs) in vitro and chromatin immunoprecipitation (ChIP)-qPCR assays in vivo indicated that FkbR1 bound to the intergenic region of fkbR1-fkbE. To investigate the roles of the target genes fkbE and fkbF in ascomycin production, the deletion and overexpressions of fkbE and fkbF were implemented, respectively. Overexpression of fkbE resulted in a 45.6% increase in ascomycin production, but little change was observed in fkbF overexpression strain. To further enhance ascomycin production, the fkbR1 and fkbE combinatorial overexpression strain OfkbRE was constructed with the ascomycin yield increased by 69.9% to 536.7 mg/L compared with that of the parent strain. Our research provided a helpful strategy to increase ascomycin production via engineering FkbR1 and its target gene.