Structural Biochemistry/Enzyme Catalytic Mechanism/Nucleoside Monophosphate Kinase

Nucleoside Monophosphate (NMP) Kinases are enzymes that aid in transferring the phosphoryl group at the end of a nucleoside triphosphate to the phosphoryl group that is on a nucleoside monophosphate. The challenge for NMP kinases is to promote the transfer of the phosphoryl group from NTP to NMP without promotoing the competing reaction - the transfer of a phosphoryl group from NTP to water; that is NTP hydrolysis. NMP kinases are known to have P-Loop Structures.



Function
Nucleoside monophosphate kinase catalyzes the reaction of ATP and NMP to yield ADP and nucleoside diphosphate (NDP). Other groups such as sugars and single-carbon groups can be substituted in this reaction instead of the phosphoryl group. One setback that may arise during this interconversion is trying not to allow the phosphoryl group from NTP to transfer to water. This can also be known as NTP hydrolysis. However, the ability of the enzyme to undergo induced fit—changing the structure in order to bind to a substrate—allows this reaction to occur successfully and bind to nucleotides instead of water. Therefore, NMP kinases is an example of catalysis by approximation. NMP kinases can exist in two forms: (1) free, or (2) bounded to substrates. NMP kinases are homologous proteins that have a conserved NTP-binding domain. This domain consists of a central beta sheet surrounded on both sides by alpha helices. NMP kinases are considered to be in P-loops because there is a loop between the first helix and the first beta strand, often in the animo sequence of X-X-X-X-Gly-Lys. The P-loop is called so because it is known to react with the beta phosphoryl group attached on the nucleotide.

Magnesium or complexes of NTP's are the true substrates for this reaction. The enzymes are essentially inactive in the absence of divalent metal ions such as the ones mentioned above. Nucleotides such as ATP bind these ions and it is the metal ion-nucleotide complex that is the true substrate for the enzymes. How does binding of a Mg or Mn ion to the nucleotide enhance catalysis? The interaction between the magnesium ion and the phosphoryl group oxygen atoms hold the nucleotide in a well-defined conformation that can be bound by an enzyme in a specific way. Magnesium ions are usually coordinated to six group in an octahedral arrangement. Typically, two oxygen atoms are directly corrdinated to a magnesium ion, with the remaining four positions often occupied by water molecules. Oxygen atoms of the alpha and beta, or the beta and gama, or alpha and gama phosphoryl groups may contribute, depending on the particular enzyme. Thus the magnesium ion provides additional points of interaction between the ATP-magnesium ion complex and the enzyme, increasing the binding energy.

ATP binding induces large conformational changes. For instance in the case of adenylate kinase the presence of the ATP substrate induces large structural changes in the kinase. This interaction caused the p-loop to be brought down onto the ATP to in order to interact with the beta phosphoryl group.The movement of the P-loop brings down the top domain of the enzyme to form a lid over the bound nucleotide. The ATP is held in position by the lid with the gama phorsphoryl group positioned next to the binding site for the second substrate NMP. The binding of NMP induces additional confomational changes. Both sets of changes ensure that a catalytically competent conformation is formed only when both the donor and the acceptor are bound, oreventing wasteful transfer of the phosphoryl group to water.

Another feature of NMP kinases, is that it interacts with the ATP substrate only after it forms a complex. ATP forms a complex with either a magnesium or manganese ion which provides more points for the substrate and enzyme to interact thus increasing the binding energy. There are isomeric forms to the metal ion-nucleotide complex depending on the interaction between the metal ion and the oxygen atoms attached to the phosphoryl groups.