The present disclosure relates to a novel protein variant having an activity of exporting 5′-inosine monophosphate, a microorganism comprising the protein variant, and a method for preparing 5′-inosine monophosphate using the microorganism.

The present disclosure relates to a novel protein variant having an activity of exporting 5′-inosine monophosphate, a microorganism comprising the protein variant, and a method for preparing 5′-inosine monophosphate using the microorganism.

An acetyl-CoA producing microorganism obtained by imparting at least one enzymatic activity selected from the group consisting of malate thiokinase, malyl-CoA lyase, glyoxylate carboligase, 2-hydroxy-3-oxopropionate reductase, and hydroxypyruvate reductase, to a microorganism that does not have any of the following (a), (b), (c), (d) or (e): (a) a carbon dioxide fixation cycle including an enzymatic reaction from malonyl-CoA to malonate semialdehyde or 3-hydroxypropionate; (b) a carbon dioxide fixation cycle including an enzymatic reaction from acetyl-CoA and CO.sub.2 to pyruvate; (c) a carbon dioxide fixation cycle including an enzymatic reaction from crotonyl-CoA and CO.sub.2 to ethylmalonyl-CoA or glutaconyl-CoA; (d) a carbon dioxide fixation cycle including an enzymatic reaction from CO.sub.2 to formate; or (e) at least one selected from the group consisting of malate thiokinase and malyl-CoA lyase.

An acetyl-CoA producing microorganism obtained by imparting at least one enzymatic activity selected from the group consisting of malate thiokinase, malyl-CoA lyase, glyoxylate carboligase, 2-hydroxy-3-oxopropionate reductase, and hydroxypyruvate reductase, to a microorganism that does not have any of the following (a), (b), (c), (d) or (e): (a) a carbon dioxide fixation cycle including an enzymatic reaction from malonyl-CoA to malonate semialdehyde or 3-hydroxypropionate; (b) a carbon dioxide fixation cycle including an enzymatic reaction from acetyl-CoA and CO.sub.2 to pyruvate; (c) a carbon dioxide fixation cycle including an enzymatic reaction from crotonyl-CoA and CO.sub.2 to ethylmalonyl-CoA or glutaconyl-CoA; (d) a carbon dioxide fixation cycle including an enzymatic reaction from CO.sub.2 to formate; or (e) at least one selected from the group consisting of malate thiokinase and malyl-CoA lyase.

The present invention relates to a process for the multi-step enzymatic conversion of adenosine diphosphate (ADP) to adenosine triphosphate (ATP), the process comprising the steps of: a) enzyme-catalyzed conversion of adenosine diphosphate in the presence of sucrose and a sucrose synthase to adenosine diphosphate-glucose; and b) enzyme-catalyzed conversion of the adenosine diphosphate-glucose formed in process step a) in the presence of inorganic pyrophosphate and a pyrophosphorylase to adenosine triphosphate and glucose-1-phosphate. Furthermore, the invention relates to the use of the process for the preparation of sugar phosphates, nucleotide sugars, glycans, glycoproteins, glycolipids or glycosaminoglycans.

The present invention relates to a process for the multi-step enzymatic conversion of adenosine diphosphate (ADP) to adenosine triphosphate (ATP), the process comprising the steps of: a) enzyme-catalyzed conversion of adenosine diphosphate in the presence of sucrose and a sucrose synthase to adenosine diphosphate-glucose; and b) enzyme-catalyzed conversion of the adenosine diphosphate-glucose formed in process step a) in the presence of inorganic pyrophosphate and a pyrophosphorylase to adenosine triphosphate and glucose-1-phosphate. Furthermore, the invention relates to the use of the process for the preparation of sugar phosphates, nucleotide sugars, glycans, glycoproteins, glycolipids or glycosaminoglycans.

The present invention relates to efficient processes useful in the preparation of fluorinated nucleosides, such as (2S,3R,4S,5R)-5-(2-amino-6-oxo-1,6-dihydro-9H-purin-9-yl)-3-fluoro-4-hydroxy-2-(mercaptomethyl)tetrahydrofuran-3-yl dihydrogen phosphate, also known as 3′-fluoro-thio-guanosine monophosphate or 3′-F-thio-GMP. Such fluorinated nucleosides may be useful as a biologically active compound and or as an intermediate for the synthesis of more complex biologically active compounds. The present invention also encompasses intermediates useful in the disclosed synthetic processes and the methods of their preparation. (I)

##STR00001##

The present invention relates to efficient processes useful in the preparation of fluorinated nucleosides, such as (2S,3R,4S,5R)-5-(2-amino-6-oxo-1,6-dihydro-9H-purin-9-yl)-3-fluoro-4-hydroxy-2-(mercaptomethyl)tetrahydrofuran-3-yl dihydrogen phosphate, also known as 3′-fluoro-thio-guanosine monophosphate or 3′-F-thio-GMP. Such fluorinated nucleosides may be useful as a biologically active compound and or as an intermediate for the synthesis of more complex biologically active compounds. The present invention also encompasses intermediates useful in the disclosed synthetic processes and the methods of their preparation. (I)

##STR00001##

The invention provides a method for producing methacrylyl-CoA that converts 3-hydroxyisobutyryl-CoA into methacrylyl-CoA using an enzyme having dehydratase activity as a method for producing methacrylyl-CoA using an enzyme catalyst. In this production method, conversion rate of 3-hydroxyisobutyryl-CoA into methacrylyl-CoA by the enzyme having dehydratase activity is 50% or higher. In this production method, furthermore, the enzyme having dehydratase activity derives from a microorganism belonging to the genus Pseudomonas or Rhodococcus.

The invention provides a method for producing methacrylyl-CoA that converts 3-hydroxyisobutyryl-CoA into methacrylyl-CoA using an enzyme having dehydratase activity as a method for producing methacrylyl-CoA using an enzyme catalyst. In this production method, conversion rate of 3-hydroxyisobutyryl-CoA into methacrylyl-CoA by the enzyme having dehydratase activity is 50% or higher. In this production method, furthermore, the enzyme having dehydratase activity derives from a microorganism belonging to the genus Pseudomonas or Rhodococcus.