Enzymatic properties and substrate specificity of a bacterial phosphatidylcholine synthase

Aktas, Meriyem; Köster, Stefan; Kizilirmak, Sarah; Casanova, Javier Carrera; Betz, Heidi; Fritz, Christiane; Moser, Roman; Yıldız, Özkan; Narberhaus, Franz

doi:10.1111/febs.12877

Cited by 16 publications

(12 citation statements)

References 61 publications

(88 reference statements)

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“…The observation that CDP-DAG binding is a prerequisite for inositol phosphate binding (and hence catalysis) implies that Mt PIPS follows a sequential ordered bi-bi reaction mechanism in which CDP-DAG binds first, followed by inositol phosphate, and the likely formation of a reactive phosphoryl intermediate through the action of an aspartate residue (D 4 in the signature sequence) acting as a catalytic base, in this case D93. This is consistent with mechanisms described previously for several other members of the family 7 21 22 23 .…”

Section: Discussionsupporting

confidence: 93%

Structural basis for phosphatidylinositol-phosphate biosynthesis

et al. 2015

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Phosphatidylinositol is critical for intracellular signalling and anchoring of carbohydrates and proteins to outer cellular membranes. The defining step in phosphatidylinositol biosynthesis is catalysed by CDP-alcohol phosphotransferases, transmembrane enzymes that use CDP-diacylglycerol as donor substrate for this reaction, and either inositol in eukaryotes or inositol phosphate in prokaryotes as the acceptor alcohol. Here we report the structures of a related enzyme, the phosphatidylinositol-phosphate synthase from Renibacterium salmoninarum, with and without bound CDP-diacylglycerol to 3.6 and 2.5 Å resolution, respectively. These structures reveal the location of the acceptor site, and the molecular determinants of substrate specificity and catalysis. Functional characterization of the 40%-identical ortholog from Mycobacterium tuberculosis, a potential target for the development of novel anti-tuberculosis drugs, supports the proposed mechanism of substrate binding and catalysis. This work therefore provides a structural and functional framework to understand the mechanism of phosphatidylinositol-phosphate biosynthesis.

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Section: Discussionsupporting

confidence: 93%

Structural basis for phosphatidylinositol-phosphate biosynthesis

et al. 2015

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“…The experimental findings presented here and in previous papers (8,13) suggest that the high-affinity Mn importer MntH is a critical virulence determinant for Brucella strains because it provides these bacteria with sufficient levels of Mn to support their PykM and SodA activities. However, further biochemical and genetic analyses are needed to gain a better understanding of the relative contributions of Mn-dependent enzymes to the virulence of Brucella strains because several other proteins that have been linked to the virulence of Brucella strains, such as the (p)ppGpp synthetase/ hydrolase Rsh (42), the phosphatidylcholine synthase Pcs (43,44), and the cyclic di-GMP phosphodiesterases BpdA and BpdB (45), are also likely to be Mn-dependent enzymes based on the biochemical properties of their counterparts in other bacteria (46)(47)(48).…”

Section: Discussionmentioning

confidence: 99%

The Manganese-Dependent Pyruvate Kinase PykM Is Required for Wild-Type Glucose Utilization by Brucella abortus 2308 and Its Virulence in C57BL/6 Mice

et al. 2018

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Pyruvate kinase plays a central role in glucose catabolism in bacteria, and efficient utilization of this hexose has been linked to the virulence of Brucella strains in mice. The brucellae produce a single pyruvate kinase which is an ortholog of the Bradyrhizobium manganese (Mn)-dependent pyruvate kinase PykM. A biochemical analysis of the Brucella pyruvate kinase and phenotypic analysis of a Brucella abortus mutant defective in high-affinity Mn import indicate that this enzyme is an authentic PykM ortholog which functions as a Mn-dependent enzyme in vivo. The loss of PykM has a negative impact on the capacity of the parental 2308 strain to utilize glucose, fructose, and galactose but not on its ability to utilize ribose, xylose, arabinose, or erythritol, and a pykM mutant displays significant attenuation in C57BL/6 mice. Although the enzyme pyruvate phosphate dikinase (PpdK) can substitute for the loss of pyruvate kinase in some bacteria and is also an important virulence determinant in Brucella, a phenotypic analysis of B. abortus 2308 and isogenic pykM, ppdK, and pykM ppdK mutants indicates that PykM and PpdK make distinctly different contributions to carbon metabolism and virulence in these bacteria. IMPORTANCE Mn plays a critical role in the physiology and virulence of Brucella strains, and the results presented here suggest that one of the important roles that the high-affinity Mn importer MntH plays in the pathogenesis of these strains is supporting the function of the Mn-dependent kinase PykM. A better understanding of how the brucellae adapt their physiology and metabolism to sustain their intracellular persistence in host macrophages will provide knowledge that can be used to design improved strategies for preventing and treating brucellosis, a disease that has a significant impact on both the veterinary and public health communities worldwide. Citation Pitzer JE, Zeczycki TN, Baumgartner JE, Martin DW, Roop RM, II. 2018. The manganesedependent pyruvate kinase PykM is required for wild-type glucose utilization by Brucella abortus 2308 and its virulence in C57BL/6 mice. J Bacteriol 200:e00471-18. https://doi.org/10 .1128/JB. Downloaded fromin vitro cultivation (5), but for others, this metal only appears to be required when the bacteria are subjected to an environmental stress, such as an exposure to reactive oxygen species (ROS) (6). Mn-dependent superoxide dismutases such as SodA, for instance, are important cytoplasmic antioxidants in bacteria (7), and in some cases, bacteria can substitute Mn for Fe in cellular proteins that do not require the redox activity of the latter metal for their function as a means of protecting these proteins from damage mediated by ROS via Fenton chemistry (6).Brucella strains produce a single high-affinity Mn transporter, MntH, and a phenotypic analysis of an isogenic mntH mutant derived from B. abortus 2308 indicates that Mn plays an important role in the basic physiology of these bacteria (8). The B. abortus mntH mutant, for instance, displays delayed growth compared to...

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“…The soybean symbiont Bradyrhizobium japonicum produces PC by the combined action of two Pmt enzymes (PmtA and PmtX1) (Minder et al, 2001;Hacker et al, 2008). In an alternative pathway, a PC synthase (Pcs) enzyme produces PC by condensation of choline and CDP-DAG (Sohlenkamp et al, 2000;Aktas et al, 2014b). Some bacteria encode only one of the two pathways, whereas other species use both pathways simultaneously (Geiger et al, 2013).…”

Section: Introductionmentioning

confidence: 99%