The present invention relates to an enzyme-catalyzed process for producing GDP-fucose from low-cost substrates guanosine and L-fucose or guanosine and D-Mannose in a single reaction mixture. Said process can be operated (semi)continuously or in batch mode. Further, said process can be adapted to produce fucosylated molecules and biomolecules including glycans, such as human milk oligosaccharides, proteins, peptides, glycoproteins or glycopeptides.

The present invention includes novel systems, methods, and compositions for the enzymatic/chemical production of pseudouridine (?) and its variants, such as N1-methyl-pseudouridine-5-triphosphate (m1?TP).

The invention relates to an in vitro cell-free expression system incorporating a novel inorganic polyphosphate-based energy regeneration system. In certain embodiments, the invention includes a cell-free expression system where the cellular energy source, ATP, is regenerated from inorganic polyphosphate using a dual enzyme system. In this embodiment, this dual enzyme system may include thermostable Adenosyl Kinase, and/or Polyphosphate Kinase enzymes.

The present disclosure provides enzymatic methods for the production of 3-phosphate-nucleoside-5-triphosphate (NQP). The present disclosure further provides enzymatic methods for the production of nucleotide-5-triphosphates.

Geranyl pyrophosphate (GPP) is a key intermediate molecule in the bioproduction of thousands of natural products. Currently, natural products are either cultivated from plants, synthesized via complex chemical synthesis strategies, or through cell-based factories also known as biofoundries. However, in order to replicate the process in a cell free environment, numerous enzymes and cofactors must be utilized making this approach costly and unviable. In order to make this process viable, a new approach was needed that uses fewer enzymes and co-factors. As described herein, the present invention demonstrates that it is possible to create GPP from glycerol through a short and concise biosynthetic pathway outside of the cell.

The present invention discloses a production method of enzymatic reaction using adenosine instead of ATP. The method comprises the following steps: (1) adding ATP regeneration enzyme, AK enzyme and adenosine in proportion to carry out an enzymatic reaction in an enzymatic reaction system; (2) separating the ATP regeneration enzyme and AK enzyme by either directly separating ATP regeneration enzyme and AK enzyme immobilized in a reaction tank, or separating free ATP regeneration enzyme and AK enzyme by an ultrafiltration membrane in a filter; and (3) separating and purifying the filtrate of step (2) to obtain a product. The disclosed method provides: greatly reduced industrial production costs; faster reaction rate; stable enzyme recovery system that is energy efficient and environmentally friendly; and capability of reusing the byproducts or collecting them for the production of ATP.

The present invention includes novel systems, methods, and compositions for the enzymatic/chemical production of pseudouridine (?) and its variants, such as N1-methyl-pseudouridine-5-triphosphate (m1?TP).

Provided is a microorganism of the genus Komagataeibacter having enhanced cellulose productivity and yield, a method of producing cellulose by using the same, and a method of producing the microorganism.

The present invention provides a coupled enzyme system, comprises: a first enzyme, comprising a polyphenol phosphorylation synthetase; a second enzyme, which is ATP regeneration enzyme; and a substrate, being phosphorylated by the first enzyme. The coupled enzyme system of the present invention integrates polyphenol phosphorylation synthetase with ATP regeneration enzyme so that the polyphenol phosphorylation synthetase is used to phosphorylate polyphenol and the ATP regeneration enzyme regenerate ATP from AMP. Therefore, the present invention not only improves the water-solubility and bioavailability of the phenolic phytochemicals but also significantly reduces ATP consumption, presenting the potential of enzymatic systems in the production of polyphenol monophosphates.

The present invention relates to a method for producing cytidine 5-monophospho-N-acetyl-neuraminic acid (CMP-Neu5Ac, 1) from low-cost substrates N-acetyl-D-glucosamine (GlcNAc), pyruvate, cytidine and polyphosphate in a single reaction mixture with a set of optionally immobilized or optionally co-immobilized enzymes comprising N-acylglucoamine 2-epimerase (AGE), an N-acetylneuraminate lyase (NAL), an N-acylneuraminate cytidylyltransferase (CSS), a uridine kinase (UDK), a uridine monophosphate kinase and a polyphosphate kinase 3 (PPK3). Further, said process may be adapted to produce Neu5Acylated i.e. sialylated biomolecules and biomolecules including a saccharide, a peptide, a protein, a glycopeptide, a glycoprotein, a glycolipid, a glycan, an antibody, and a glycoconjugate, in particular, an antibody drug conjugate, and a carbohydrate conjugate vaccine, or a flavonoid.