New Crystal Structure Helps Explain 50 Years of Research on a Novel Membrane
Enzyme in E. coli
Diacylglycerol kinase is an integral membrane enzyme involved in the synthesis of key
components of the cellular envelope, including lipopolysaccharide, in Gram-negative
bacteria. For 50 years, it has served as a paradigm for investigating membrane protein
enzymology, folding, assembly, and stability. With a view to understanding how this enzyme
functions at a molecular level, the crystal structure of diacylglycerol kinase from Escherichia
coli has been determined at high resolution using state-of-the-art X-ray crystallographic
techniques and X-ray synchrotron sources. The structure rationalizes extensive published
biochemical and biophysical data on this novel kinase.
Diacylglycerol kinase is an enzyme that catalyzes a complex reaction involving substrates
with widely contrasting solubilities. Its lipid substrate abhors water and resides in the fatty
membrane that surrounds the cell. By contrast its second or co-substrate, ATP (which
incidentally is the energy currency of the cell), is totally water soluble. How this diminutive
enzyme, the smallest known kinase, manages to bring such disparate substrates together at
the membrane interface for reaction is revealed in molecular detail in the new crystal
structure. The protein embeds most of its bulk in the membrane where it interacts with the
lipid substrate. The rest extends out of the membrane into the watery cytoplasm to sequester
ATP. The two substrates are then forced into a molecular embrace at the membranecytoplasm interface. Residing just atomic distances apart and perfectly aligned for reaction in
the maw of the enzyme the two molecules are thought to fuse becoming a single entity, ever
so briefly. Upon separating, the lipid takes with it ATP’s terminal phosphate. In a series of
follow up enzyme reactions, the newly formed phospholipid contributes to cell envelop
synthesis.
These findings are likely to attract considerable attention, in part, because they help explain
almost half a century of work on this paradigmatic membrane protein. The study is notable
too because the crystal structure differs profoundly from the only other structure available for
this novel kinase that was obtained using solution nuclear magnetic resonance (see News &
Views).
The research was carried out in Professor Martin Caffrey’s Membrane Structural and
Functional Biology (MS&FB) group in Trinity’s School of Medicine and School of
Biochemistry and Immunology. The first author on the paper, Dr. Dianfan Li, a Research
Associate in the MS&FB group, led the 4-year-long project that involved state-of-the-art
recombinant DNA technology and membrane protein production, purification, and
crystallization methods. The structure was solved based on diffraction measurements
performed using highly specialized X-ray beams at synchrotron radiation facilities in the
USA (Advanced Photon Source) and the UK (Diamond Light Source). The work was funded
through grants from Science Foundation Ireland and the US National Institutes of Health.
Li, D., Lyons, A.J., Pye, V.E., Vogeley, L., Aragao, D., Kenyon, C.P., Shah, S.T.A., Doherty, C.,
Aherne, M., Caffrey, M. (2013) Crystal structure of the integral membrane diacylglycerol kinase.
Nature 49: 521-524. DOI 10.1038/nature12179 http://dx.doi.org/10.1038/nature12179
News & Views: http://www.nature.com/nature/journal/vaop/ncurrent/full/nature12245.html