Provided herein, in some aspects, are methods and compositions for cell-free production of ribonucleic acid.

The present disclosure is related to biosynthetic methods of forming monosaccharides, and systems for generating the same. A benefit of the methods and systems disclosed herein can include the sustainable production of monosaccharides in an automated process. A benefit of the methods and systems herein can be the generation of monosaccharides from renewable source materials. An additional benefit of the methods and systems herein can include the use of abundant feedstocks, such as carbon dioxide, for the efficient generation of select monosaccharides for use as nutrients and for other useful applications. Another benefit of the methods and systems disclosed herein can include reduction of excess carbon dioxide from the environment.

The present disclosure is related to electrochemical methods of forming monosaccharides, and systems for generating the same. A benefit of the methods and systems disclosed herein can include the sustainable production of monosaccharides in an automated process. A benefit of the methods and systems herein can be the generation of monosaccharides from renewable source materials. An additional benefit of the methods and systems herein can include the use of abundant feedstocks, such as carbon dioxide, for the efficient generation of select monosaccharides for use as nutrients and for other useful applications. Another benefit of the methods and systems disclosed herein can include reduction of excess carbon dioxide from the environment.

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 disclosure discloses a strain and method for producing rosmarinic acid, and belongs to the technical field of bioengineering. The disclosure constructs a recombinant cell or a combination of recombinant cells expressing 4-coumarate: CoA ligase, rosmarinic acid synthase, polyphosphate kinase 2-I (PPK2-I) and polyphosphate kinase 2-II (PPK2-II), and utilizes the recombinant cell or the combination of recombinant cells to catalyze Danshensu and caffeic acid for synthesizing rosmarinic acid. The disclosure has good industrial application prospects.

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.

An object of the present invention is to provide a method for producing nicotinamide mononucleotide (NMN) with excellent production efficiency. The method for producing NMN according to the present invention (the first aspect) comprises the step of bringing a transformant with enhanced expression of enzymes nicotinamide phosphoribosyltransferase (Nampt), phosphoribosyl pyrophosphate synthetase (Prs) and polyphosphate kinase (Ppk), a cell-free protein synthesis reaction solution having the three enzymes expressed, or a treated product thereof into contact with a mixture containing ribose-5-phosphate (R5P), nicotinamide (NAM), ATP and polyphosphate.

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.

A method of producing a substance includes synthesizing a molecule at least by mixing substrates, a synthase, adenosine triphosphate (ATP), a polyphosphate kinase 2, and a polyphosphoric acid mixture. The polyphosphoric acid mixture includes 50% or more of polyphosphoric acid with a degree of polymerization of not less than 15. Adenosine diphosphate (ADP) is generated from the ATP during the synthesis. The synthesis is coupled with an ATP regeneration reaction in which the ATP is regenerated by the polyphosphate kinase 2 from the ADP and the polyphosphoric acid.

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.