Structural Insights into Broad-Range Polyphosphate Kinase 2-II Enzymes Applicable for Pyrimidine Nucleoside...

Area of Science:

• Biochemistry
• Enzymology
• Molecular Biology

Background:

• Polyphosphate kinases (PPK) are vital enzymes involved in energy metabolism and cellular stress responses.
• PPK family 2 (PPK2) enzymes specifically utilize inorganic polyphosphate (polyP) to phosphorylate nucleotides.

Purpose of the Study:

• To characterize Bacillus cereus PPK2 (BcPPK2) and Lysinibacillus fusiformis PPK2 (LfPPK2) enzymes.
• To investigate their substrate specificity and potential for selective nucleotide synthesis.

Main Methods:

• Enzyme characterization of BcPPK2 and LfPPK2.
• Analysis of substrate specificity for pyrimidine and purine nucleoside monophosphates (NMPs).
• Preparative enzymatic synthesis and structural analysis (crystal structures) of PPK2 enzymes.

Main Results:

• Both BcPPK2 and LfPPK2 efficiently converted over 85% of pyrimidine NMPs to nucleoside diphosphates (NDPs).
• NTP formation was specific to purine NMPs.
• Successful preparative synthesis of cytidine diphosphate (CDP) with a 49% yield.
• Structural insights into the active sites and substrate interactions of BcPPK2 and LfPPK2.

Conclusions:

• PPK2-II enzymes are effective biocatalysts for the selective synthesis of pyrimidine NDPs.
• The findings provide a basis for utilizing these enzymes in biotechnological applications for nucleotide production.