4DA5

Choline Kinase alpha acts through a double-displacement kinetic mechanism involving enzyme isomerisation, as determined through enzyme and inhibitor kinetics and structural biology

Summary for 4DA5

• Entry DOI: 10.2210/pdb4da5/pdb
• Descriptor: Choline kinase alpha, SULFATE ION, (3R)-1-azabicyclo[2.2.2]oct-3-yl[bis(5-chlorothiophen-2-yl)]methanol, ... (4 entities in total)
• Functional Keywords: kinase, signal transduction, cytoplasmic, transferase-transferase inhibitor complex, transferase/transferase inhibitor
• Biological source: Homo sapiens (human)
• Cellular location: Cytoplasm: P35790
• Total number of polymer chains: 2
• Total formula weight: 105562.56

Authors

Brown, K.,Hudson, C.,Charlton, P.,Pollard, J. (deposition date: 2012-01-12, release date: 2013-04-17, Last modification date: 2024-02-28)

Primary citation

Hudson, C.S.,Knegtel, R.M.,Brown, K.,Charlton, P.A.,Pollard, J.R.
Kinetic and mechanistic characterisation of Choline Kinase-alpha.
Biochim.Biophys.Acta, 1834:1107-1116, 2013

PubMed Abstract: Choline Kinase is a key component of the Kennedy pathway that converts choline into a number of structural and signalling lipids that are essential for cell growth and survival. One member of the family, Choline Kinase-α (ChoKα) is frequently up-regulated in human cancers, and expression of ChoKα is sufficient to transform cells. Consequently ChoKα has been studied as a potential target for therapeutic agents in cancer research. Despite great interest in the enzyme, mechanistic studies have not been reported. In this study, a combination of initial velocity and product inhibition studies, together with the kinetic and structural characterisation of a novel ChoKα inhibitor is used to support a mechanism of action for human ChoKα. Substrate and inhibition kinetics are consistent with an iso double displacement mechanism, in which the γ-phosphate from ATP is transferred to choline in two distinct steps via a phospho-enzyme intermediate. Co-crystal structures, and existing site-specific mutation studies, support an important role for Asp306, in stabilising the phospho-enzyme intermediate. The kinetics also indicate a distinct kinetic (isomerisation) step associated with product release, which may be attributed to a conformational change in the protein to disrupt an interaction between Asp306 and the phosphocholine product, facilitating product release. This study describes a mechanism for ChoKα that is unusual amongst kinases, and highlights the availability of different enzyme states that can be exploited for drug discovery.

PubMed: 23416529
DOI: 10.1016/j.bbapap.2013.02.008

Experimental method

X-RAY DIFFRACTION (2.4 Å)