Uridine Kinase

Thomas Traut

Research Interests

Dissociating Enzymes

UMP Synthase

Uridine Kinase

Beta-alanine Synthase

Purine & Pyrimidine Concentrations

This enzyme is ubiquitous in all mammalian cells/tissues, and important in most micro-organisms. It is allosterically regulated between the active tetramer and inactive monomer by CTP and ATP. 



Sequence: the mammalian cDNA, as well as 4 microorganisms, have recently been sequenced. This provided the location of 3 polypeptide sequences identified with the ATP, or phosphate donor site (shown in yellow), and also 3 peptide sequences identified with the acceptor substrate site (green).


The position of peptide segments associated with the phosphate donor site (usually ATP) and the phosphate acceptor site (uridine or cytidine) is shown with a few specific sequence motifs in upper diagram.  The lower diagram displays more correctly the size and general position of these peptide segments.  Most kinases have the three Kinase motifs, and a few  nucleoside kinases and nucleotide kinases have the same pattern of acceptor motifs.

Alignment of uridine kinase from mouse, with sequences for three enzymes for which a crystal structure shows the amino acids that contact the bound ligands, which are highlighted in red.  The same residues in uridine kinase, at these positions are shown in green; some consensus residues for deoxy-thymidine kinase at the Kinase-1 and Kinase-3 sites are in orange.



Important Question: how is regulation achieved?


Is there a regulatory site for CTP, or ATP? Or, does CTP act by binding backwards in the catalytic site as a bi-substrate analog? Deoxycytidine is also a poor substrate, at the phosphate acceptor site.  It is then interesting that dCTP acts as a modest substrate (P-donor) and does not dissociate the enzyme tetramer.  In contrast, CTP only inhibits and causes the tetramer to dissociate.


With a mutant of the mouse uridine kinase that had a single replacement at position 146, kinetic studies showed a similar loss in binding affinity for the substrate uridine, and for the inhibitor CTP.  Various kinetic studies were consistent with CTP binding "backwards" at the catalytic site as a bisubstrate analog.




This figure illustrates how CTP binds "backwards" at the catalytic site as a physiological effector of uridine kinase.  The upper panel shows Cytidine at the Acceptor site and ATP at the phosphate donor site.  The base (Ad) at the donor site presumably has little or no interaction with the enzyme, since purines and pyrimidine nucleoside triphosphates (NTPs) may be  P-donors. The lower panel shows the inhibibitor CTP, binding backwards compared to other NTPs that are P-donors. This synergism in binding at two sites also accounts for the better binding affinity of CTP relative to ATP and cytidine.


Relevant Paper:

1.   Ropp, P.A. and Traut, T.W. (1998)  Uridine kinase: altered enzyme with decreased affinities for uridine and CTP.  Arch. Bioch. Biophys. 359, 63-68.