The disclosure provides biosynthetic platforms that generate olivetolic acid and its analogues at high titers from microbes, and in cell free systems.

The present disclosure provides recombinant microorganisms and methods for the anaerobic production of 2,4-furandimethanol from one or more carbon sources. The microorganisms and methods provide redox-balanced and ATP positive pathways for co-producing 2,4-furandimethanol with ethanol and for co-producing 2,4-furandimethanol with ethanol and acetone and/or isopropanol. The method provides recombinant microorganisms that express endogenous and/or exogenous nucleic acid molecules encoding polypeptides that catalyze the conversion of a carbon source into 2,4-furandimethanol and that couple the 2,4-furandimethanol pathway with an additional metabolic pathway. The present disclosure further provides enzymatic production of 2,4-furandicarboxylic acid.

The present disclosure provides recombinant microorganisms and methods for the anaerobic production of 2,4-furandimethanol from one or more carbon sources. The microorganisms and methods provide redox-balanced and ATP positive pathways for co-producing 2,4-furandimethanol with ethanol and for co-producing 2,4-furandimethanol with ethanol and acetone and/or isopropanol. The method provides recombinant microorganisms that express endogenous and/or exogenous nucleic acid molecules encoding polypeptides that catalyze the conversion of a carbon source into 2,4-furandimethanol and that couple the 2,4-furandimethanol pathway with an additional metabolic pathway. The present disclosure further provides enzymatic production of 2,4-furandicarboxylic acid.

The present disclosure provides engineered human extracellular DNASE proteins (e.g., variants of DNASE1 (D1), DNASE1-LIKE 1 (D1L1), DNASE1-LIKE 2 (DTL2), DNASE1-LIKE 3 Isoform 1 (DTL3), DNASE1-LIKE 3 Isoform 2 (DTL3-2), DNASE2A (D2A), and DNASE2B (D2B)) that are useful for treating conditions characterized by neutrophil extracellular trap (NET) accumulation and/or release. In accordance with the invention, the DNase variant has advantages for therapy and/or large-scale manufacturing.

This invention is an improved method of robust and scalable production of precursors of active cannabinoids, including geranyl pyrophosphate (GPP) and/or cannabigerolic acid (CBGA), in a methylotrophic yeast host cell. The improved methods incorporate a polypeptide encoding an Erg20 variant (F98W/N128W) into a methylotrophic yeast host cell, for example Pichia pastoris (Komagataella phaffii), that biases the natural production of FPP and GPP towards GPP, a precursor to the intermediate CBGA, crucial to the synthesis of active cannabinoids.

This invention is an improved method of robust and scalable production of precursors of active cannabinoids, including geranyl pyrophosphate (GPP) and/or cannabigerolic acid (CBGA), in a methylotrophic yeast host cell. The improved methods incorporate a polypeptide encoding an Erg20 variant (F98W/N128W) into a methylotrophic yeast host cell, for example Pichia pastoris (Komagataella phaffii), that biases the natural production of FPP and GPP towards GPP, a precursor to the intermediate CBGA, crucial to the synthesis of active cannabinoids.

The present disclosure methods for identifying binding partners using cell surface display libraries, where the cells of the library display engineered peptides on their cell surfaces for identification of peptides that bind to targets of interest. The engineered peptides are preferably expressed in the cells under conditions that provide both secretion and display of the engineered peptides on the cell surfaces, thus providing access of the engineered peptides to identify potential binding pairs. The cell libraries cab be engineered using an automated editing system that provides for one or more targeted edits per cell.

Isolated nucleic acids, expression methods, host cells, expression vectors, and DNA constructs for producing proteins, and proteins produced using the expression methods are disclosed. More specifically, nucleic acids isolated from Pichia pastons having promoter activity and expression methods, host cells, expression vectors, and DNA constructs of using the Pichia pastons promoters to produce different proteins and polypeptides are disclosed.

The invention discloses the application of a β-N-acetylhexosaminidase (HaHex74) from Haloferula sp. in the synthesis of human milk oligosaccharides. The invention provides the use of HaHex74 protein or related biological materials thereof in any one of the following: synthesizing human milk oligosaccharides; synthesizing Lacto-N-triose II and/or Lacto-N-neotetraose; the HaHex74 protein having the amino acid sequence shown in SEQ ID No. 2 is derived from Haloferula sp. The β-N-acetylhexosaminidase HaHex74 disclosed by the invention possesses high-level expression, excellent hydrolysis properties and transglycosylation activity, which may make it potentially useful in the production of human milk oligosaccharides.

The invention features compositions and methods for the increased production of mevalonate, isoprene, isoprenoid precursor molecules, and/or isoprenoids in microorganisms via the heterologous expression of the mvaE and mvaS genes from the organisms Listeria grayi DSM 20601, Enterococcus faecium, Enterococcus gallinarum EG2, and Enterococcus casseliflavus.