br Author contributions br Acknowledgements This work was su

Author contributions
Acknowledgements This work was supported by the State Fund for Fundamental Researches of Ukraine (grants F75-2018, F73-2018, F83-2018), Ukraine, grant No. 2.1.10.32-15 from National Academy of Sciences of Ukraine, Ukraine, Belarusian Republican Foundation for Fundamental Research (grant No. X 16К-057), Belarus and the Ministry of Education, Youth and Sports of CR from European Regional Development Fund-Project “Centre for Experimental Plant Biology” (No. CZ.02.1.01/0.0/0.0/16_019/0000738), Czech Republic. We kindly thank Prof. E. Ruelland for providing seed stocks of dgk mutants.
Introduction Diacylglycerol kinases (DGKs) are transferases that play essential roles in the physiology of a number of cell types (Baldanzi, 2014; Merida et al., 2017; Shulga et al., 2011a; Tu-Sekine and Raben, 2011). These enzymes catalyze the phosphorylation of diacylglycerol to generate phosphatidic PNU-120596 sale and use ATP as the phosphate donor with the exception of the yeast DGK which uses CTP (Han et al., 2008). In contrast to our understanding of the structure and catalytic mechanism of other lipid and protein kinases, understanding of DGKs is limited. This is particularly true for the mammalian DGKs as much more is known about prokaryotic DGKs. Interest in DGKs increased as it became clear that not only are they important for lipid homeostasis, they serve to modulate the relative levels of diacylglycerol (DAG) and phosphatidic acid (PtdOH) that play critical roles in a variety of signaling pathways (Eichmann and Lass, 2015; Liu et al., 2013) including neurotransmission (Raben and Barber, 2017; Tu-Sekine et al., 2015; Tu-Sekine and Raben, 2011). This highlights the need to understand the structure, regulation and catalytic mechanism of these enzymes. Indeed, lacking of the molecular and structural insight into DGKs has been a huge barrier for designing highly specific inhibitors for the DGKs to probe further roles and potential therapeutic strategies where possible.
Overall structural features of lipid kinases Despite the differences in lipid substrates, an overall structure of lipid kinases must contain: a region/residue(s) for membrane association/stabilization, a domain to bind and orient the phosphor donor (ATP/CTP), and a domain for the binding and orientation of the phosphoryl acceptor (lipid substrate). These domains are oriented in a manner to permit the catalytic reaction (Cheek et al., 2002; Fabbro et al., 2015). While we have gained valuable insights into the nucleotide binding domains, the structure of the lipid substrate binding is not well understood. Understanding the architecture of these domains and their relationship to each other in each lipid kinase will provide valuable insights into the catalytic mechanism(s) of these enzymes.
Prokaryotic DAG kinases There are two classes of prokaryotic DGKs designated as DGKA and DGKB (Van Horn and Sanders, 2012). Perhaps the most well-understood, and smallest, of these are the DGKAs. There are two types of DGKAs; one found in gram-positive bacteria and the other found in gram-negative bacteria. A hallmark of these DGKs are that they exist as homotrimeric proteins where each monomer contains three transmembrane-spanning regions (Li et al., 2013; Oxenoid et al., 2002). Interestingly, there are some important substrate and mechanistic differences between these two DGKAs. The one found in gram-positive bacteria prefers undecaprenol as a substrate while the gram-negative enzyme prefers diacylglycerol as a substrate although it also phosphorylates monoacylglycerol, as well as ceramide (Jerga et al., 2007). Mechanistically, it is interesting to note that this DGKA also show hydrolytic (ATPase) activity (Li et al., 2015).