Nucleoside phosphate regeneration system

A nucleoside phosphate regeneration system must meet several specifications to be practical. To be economical, a regeneration method must be capable of recycling the cofactor 102-106 times1391. All materials should be readily available, inexpensive, easily handled, stable under reaction conditions and compatible with the rest of the reaction system. The transfer of phosphate should be thermodynamically and... 

The use of isolated enzymes to form or cleave P-O bonds is an important application of biocatalysts. Restriction endonucleases, (deoxy)ribonucleases, DNA/ RNA-ligases, DNA-RNA-polymerases, reverse transcriptases etc. are central to modern molecular biology(1). Enzyme catalyzed phosphoryl transfer reactions have also found important applications in synthetic organic chemistry. In particular, the development of convenient cofactor regeneration systems has made possible the practical scale synthesis of carbohydrates, nucleoside phosphates, nucleoside phosphate sugars and other natural products and their analogs. This chapter gives an overview of this field of research. 

The most widely used and best developed enzyme-catalyzed phosphorylations are the ones that are coupled to ATP regeneration systems. Sugar phosphates, nucleoside phosphates and glycerides are the major classes of compounds prepared with these methods. 

Acetyl phosphate, which can be synthesized from acetic anhydride and phosphoric acid [526], is a commonly used regeneration system in conjunction with acetate kinase [527]. It is modestly stable in aqueous solution and while its phosphoryl donor potential is lower than that of PEP, it is considerably cheaper. As for pymvate kinase, acetate kinase also can accept nucleoside phosphates other than adenosine, and it is inhibited by acetate. 

The use of the phosphoenol pyruvate (PEP)/pyruvate kinase system is probably the most useful method for the regeneration of nucleoside triphosphates [523]. PEP is not only very stable towards spontaneous hydrolysis but it is also a stronger phosphorylating agent than ATP. Furthermore, nucleosides other than adenosine phosphates are also accepted by pyruvate kinase. The drawbacks of this system are the more complex synthesis of PEP [524, 525] and the fact that pyruvate kinase is inhibited by pyruvate at higher concentrations. 

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