The yeast Sir2 protein, required for transcriptional silencing, has an NAD + -dependent histone deacetylase (HDA) activity. Yeast extracts contain a HDA activity that is eliminated in strain from which SIR2 and its four homologs have been deleted. This also displayed by purified Sir2p homologous Archaeal, eubacterial, human proteins, depends completely on all species tested. NPT1 gene, encoding important synthesis enzyme, rDNA telomeric silencing contributes to of the HM loci. Null mutants...

Acetyl-coenzyme A (CoA) synthetase (Acs) is an enzyme central to metabolism in prokaryotes and eukaryotes. Acs synthesizes acetyl CoA from acetate, adenosine triphosphate, through acetyl–adenosine monophosphate (AMP) intermediate. Immunoblotting mass spectrometry analysis showed that Salmonella enterica activity posttranslationally regulated by acetylation of lysine-609. Acetylation blocks synthesis the adenylate intermediate but does not affect thioester-forming enzyme. Activation...

Acetyl-coenzyme A synthetase catalyzes the two-step synthesis of acetyl-CoA from acetate, ATP, and CoA belongs to a family adenylate-forming enzymes that generate an acyl-AMP intermediate. This includes other acyl- aryl-CoA synthetases, firefly luciferase, adenylation domains modular nonribosomal peptide synthetases. We have determined X-ray crystal structure complexed with adenosine-5'-propylphosphate CoA. The identifies binding pocket as well new conformation for members this enzyme in...

The fusion of yeast vacuoles, like other organelles, requires a Rab-family guanosine triphosphatase (Ypt7p), Rab effector and Sec1/Munc18 (SM) complex termed HOPS (homotypic vacuole protein sorting), soluble N-ethylmaleimide-sensitive factor attachment receptors (SNAREs). central 0-layer the four bundled vacuolar SNAREs wild-type three glutaminyl (Q) one arginyl (R) residues for optimal fusion. Alterations this layer dramatically increase K(m) value to assemble trans-SNARE complexes fuse. We...

Abstract SIR2 proteins have NAD+-dependent histone deacetylase activity, but no metabolic role has been assigned to any of these proteins. In Salmonella enterica, function was required for activity the acetyl-CoA synthetase (Acs) enzyme. A greater than two orders magnitude increase in specific Acs enzyme synthesized by a sirtuin-deficient strain measured after treatment with homogeneous S. enterica protein. Human SIR2A and yeast restored growth SIR2-deficient on acetate propionate,...

Although concentrated soluble N-ethylmaleimide-sensitive factor attachment protein receptors (SNAREs) drive liposome fusion and lysis, the of intracellular membranes also requires Rab GTPases, effectors, SM proteins, specific regulatory lipids is accompanied by little or no lysis. To rationalize these findings, we generated yeast strains that overexpress all four vacuolar SNAREs (4SNARE(++)). vacuoles with physiological levels Rab, effector/SM complex, support rapid without Rab-...

Vacuolar-proton ATPases (V-ATPases) are conserved complexes that couple the hydrolysis of ATP to pumping protons across membranes. V-ATPases known play diverse roles in cellular physiology. We studied Toxoplasma gondii V-ATPase complex and discovered a dual role pump protecting parasites against ionic stress maturation secretory proteins endosomal-like compartments. subunits localize plasma membrane acidic vesicles, characterization conditional mutants a1 subunit highlighted functionality at...

Defects in normal autophagic pathways are implicated numerous human diseases—such as neurodegenerative diseases, cancer, and cardiomyopathy—highlighting the importance of autophagy its proper regulation. Herein we show that Vibrio parahaemolyticus uses type III effector VopQ (Vibrio outer protein Q) to alter flux by manipulating partitioning small molecules ions lysosome. This binds conserved V o domain vacuolar-type H + -ATPase causes deacidification lysosomes within minutes entering host...

The studies reported here identify propionyl coenzyme A (propionyl-CoA) as the common intermediate in 1,2-propanediol and propionate catabolic pathways of Salmonella enterica serovar Typhimurium LT2. Growth on a carbon energy source led to formation excretion propionate, whose activation propionyl-CoA relied activities kinase (PduW)/phosphotransacetylase (Pta) enzyme system CobB sirtuin-controlled acetyl-CoA (Acs, PrpE) synthetases. different affinities these systems for ensure sufficient...

Posttranslational regulation of protein function by acetylation is present throughout nature. Regulation Sir2 (sirtuin) deacetylases conserved in all domains life. In the prokaryote Salmonella enterica, metabolic enzyme acetyl-coenzyme A synthetase (Acs) regulated a Sir2-dependent acetylation/deacetylation system (SDPADS). The recent identification acetyltransferase responsible for Acs defined SDPADS prokaryotes. This report identifies one residue Acs, Leu-641, which critical enzyme. vivo...

Brugia malayi, a parasitic roundworm of humans, is colonized by the obligate intracellular bacterium, Wolbachia pipientis. The symbiosis between this nematode and bacterium essential for reproduction long-term survival in human host. Therefore, identifying molecular mechanisms required to persist colonize B. malayi tissues will provide new information regarding basic biology endosymbiosis. utilize Type IV secretion system translocate so-called "effector" proteins into cytosol cells promote...

Significance Fusion of intracellular membranes is involved in many critical cellular processes, such as neurotransmission, protein trafficking, and the lysosomal degradation invading bacterial pathogens. Accordingly, some pathogens use effectors to alter host membrane fusion directly a survival mechanism. In this study, we show that Vibrio secreted effector, VopQ, potent inhibitor yeast homotypic vacuole vitro. Although VopQ was shown deacidify vacuoles via its known V-type H + -ATPase...

During infection, the intracellular pathogenic bacterium Legionella pneumophila causes an extensive remodeling of host membrane trafficking pathways, both in construction a replication-competent vacuole comprised ER-derived vesicles and plasma components, inhibition normal phagosome:endosome/lysosome fusion pathways. Here, we identify LegC3 secreted effector protein from L. as able to inhibit SNARE- Rab GTPase-dependent pathway vitro, homotypic yeast vacuoles (lysosomes). This appeared be...

Biological membrane fusion employs divalent cations as protein cofactors or signaling ligands. For example, Mg2+ is a cofactor for the N-ethylmaleimide-sensitive factor (NSF) ATPase, and Ca2+ signal from neuronal depolarization required synaptotagmin activation. Divalent also regulate liposome fusion, but role of such ion interactions with lipid bilayers in Rab- soluble NSF attachment receptor (SNARE)-dependent biological less clear. Yeast vacuole requires Sec18p ATPase activity, docking...

This report shows that Salmonella enterica catabolizes ethanolamine to acetyl-CoA (Ac-CoA), which enters the glyoxylate bypass and tricarboxylic acid cycle for generation of energy central metabolites. During growth on ethanolamine, S. excreted acetate, whose recapture depended Ac-CoA synthetase (Acs) housekeeping phosphotransacetylase (Pta) enzyme activities. The Pta did not play a role in acetate excretion during ethanolamine. It is proposed necessary maintain pool free CoA. Acetate...

Wolbachia is an unculturable, intracellular bacterium that persists within extremely broad range of arthropod and parasitic nematode hosts, where it transmitted maternally to offspring via vertical transmission. In the filarial Brugia malayi, a causative agent human lymphatic filariasis, endosymbiont, its presence essential for proper development, survival, pathogenesis. While elucidation Wolbachia:nematode interactions promote bacterium's persistence great importance, research has been...

Summary Rhodococcus equi is a multihost, facultative intracellular bacterial pathogen that primarily causes pneumonia in foals less than six months age and immunocompromised people. Previous studies determined the major virulence determinant of R. surface bound associated protein A (VapA). The presence VapA inhibits maturation ‐containing phagosomes promotes survival, as by inability vapA deletion mutants to replicate host macrophages. While mechanism action remains elusive, we show soluble...

The intracellular pathogen, Legionella pneumophila, relies on numerous secreted effector proteins to manipulate host endomembrane trafficking events during pathogenesis, thereby preventing fusion of the bacteria-laden phagosome with endolysosomal compartments, and thus escaping degradation. Upon expression in surrogate eukaryotic model Saccharomyces cerevisiae, we find that L. pneumophila LegC7/YlfA protein disrupts delivery both biosynthetic endocytic cargo yeast vacuole. We demonstrate...

Vibrio parahemolyticus Type III effector VopQ is both necessary and sufficient to induce autophagy within one hour of infection. We demonstrated that interacts with the Vo domain conserved vacuolar H(+)-ATPase. Membrane-associated subsequently forms pores in membranes acidic compartments, resulting immediate release protons without concomitant lumenal protein contents. These studies show how a bacterial pathogen can compromise host ion potentials using gated pore-forming equilibrate levels...

FlhF and FlhG control the location number of flagella, respectively, in many polar-flagellated bacteria. The roles are not well characterized bacteria that have multiple polar such as