scholarly journals Uptake of N-acetylneuraminic acid by Escherichia coli K-235. Biochemical characterization of the transport system

1987 ◽  
Vol 246 (2) ◽  
pp. 287-294 ◽  
Author(s):  
L B Rodríguez-Aparicio ◽  
A Reglero ◽  
J M Luengo
Keyword(s):  
Escherichia Coli ◽  
Sialic Acid ◽  
Transport System ◽  
Biochemical Characterization ◽  
Osmotic Shock ◽  
Metabolic Inhibitors ◽  
Acid Uptake ◽  
Uptake System ◽  
Acid Transport ◽  
Acetylneuraminic Acid

Kinetic measurement of the uptake of N-acetyl[4,5,6,7,8,9-14C]neuraminic acid by Escherichia coli K-235 was carried out in vivo at 37 degrees C in 0.1 M-Tris/maleate buffer, pH 7.0. Under these conditions uptake was linear for at least 30 min and the Km calculated for sialic acid was 30 microM. The transport system was osmotic-shock-sensitive and was strongly inhibited by uncouplers of oxidative phosphorylation [2,4-dinitrophenol (100%); NaN3 (66%]) and by the metabolic inhibitors KCN (84%) and sodium arsenate (76%). The thiol-containing compounds mercaptoethanol, glutathione, cysteine, dithiothreitol and cysteine had no significant effect on the sialic acid-transport rate, whereas the thiol-modifying reagents N-ethylmaleimide, iodoacetate and p-chloromercuribenzoate almost completely blocked (greater than 94%) the uptake of this N-acetyl-sugar. N-Acetylglucosamine inhibited non-competitively the transport of N-acetylneuraminic acid, whereas other carbohydrates (hexoses, pentoses, hexitols, hexuronic acids, disaccharides, trisaccharides) and N-acetyl-sugars or amino acid derivatives (N-acetylmannosamine, N-acetylcysteine, N-acetylproline and N-acetylglutamic acid) did not have any effect. Surprisingly, L-methionine and its non-sulphur analogue L-norleucine partially blocked the transport of this sugar (50%), whereas D-methionine, D-norleucine, several L-methionine derivatives (L-methionine methyl ester, L-methionine ethyl ester, L-methionine sulphoxide) and other amino acids did not affect sialic acid uptake. The N-acetylneuraminic acid-transport system is induced by sialic acid and is strictly regulated by the carbon source used for E. coli growth, arabinose, lactose, glucose, fructose and glucosamine being the carbohydrates that cause the greatest repressions in this system. Addition of cyclic AMP to the culture broth reversed the glucose effect, indicating that the N-acetylneuraminic acid-uptake system is under catabolic regulation. Protein synthesis is not needed for sialic acid transport.

scholarly journals A Novel Sialic Acid Utilization and Uptake System in the Periodontal Pathogen Tannerella forsythia

2010 ◽  
Vol 192 (9) ◽  
pp. 2285-2293 ◽  
Author(s):  
Sumita Roy ◽  
C. W. Ian Douglas ◽  
Graham P. Stafford
Keyword(s):  
Escherichia Coli ◽  
Sialic Acid ◽  
Outer Membrane ◽  
Cloning And Expression ◽  
Periodontal Pathogen ◽  
Acid Uptake ◽  
Uptake System ◽  
Tannerella Forsythia ◽  
Inner Membrane ◽  
Biofilm Growth

ABSTRACT Tannerella forsythia is a key contributor to periodontitis, but little is known of its virulence mechanisms. In this study we have investigated the role of sialic acid in biofilm growth of this periodontal pathogen. Our data show that biofilm growth of T. forsythia is stimulated by sialic acid, glycolyl sialic acid, and sialyllactose, all three of which are common sugar moieties on a range of important host glycoproteins. We have also established that growth on sialyllactose is dependent on the sialidase of T. forsythia since the sialidase inhibitor oseltamivir suppresses growth on sialyllactose. The genome of T. forsythia contains a sialic acid utilization locus, which also encodes a putative inner membrane sialic acid permease (NanT), and we have shown this is functional when it is expressed in Escherichia coli. This genomic locus also contains a putatively novel TonB-dependent outer membrane sialic acid transport system (TF0033-TF0034). In complementation studies using an Escherichia coli strain devoid of its outer membrane sialic acid transporters, the cloning and expression of the TF0033-TF0034 genes enabled an E. coli nanR nanC ompR strain to utilize sialic acid as the sole carbon and energy source. We have thus identified a novel sialic acid uptake system that couples an inner membrane permease with a TonB-dependent outer membrane transporter, and we propose to rename these novel sialic acid uptake genes nanO and nanU, respectively. Taken together, these data indicate that sialic acid is a key growth factor for this little-characterized oral pathogen and may be key to its physiology in vivo.

Characterization of l-arginine transport in adrenal cells: effect of ACTH

2006 ◽  
Vol 291 (2) ◽  
pp. E291-E297 ◽  
Author(s):  
Esteban M. Repetto ◽  
Vanesa Pannunzio ◽  
Francisco Astort ◽  
Camila Martinez Calejman ◽  
Marcos Besio Moreno ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Amino Acid ◽  
Transport System ◽  
Zona Fasciculata ◽  
Acid Uptake ◽  
Uptake System ◽  
Amino Acid Transport System ◽  
Cationic Amino Acid ◽  
Arginine Transport

Nitric oxide synthesis depends on the availability of its precursor l-arginine, which could be regulated by the presence of a specific uptake system. In the present report, the characterization of the l-arginine transport system in mouse adrenal Y1 cells was performed. l-arginine transport was mediated by the cationic/neutral amino acid transport system y+L and the cationic amino acid transporter (CAT) y+ in Y1 cells. These Na+-independent transporters were identified by their selectivity for neutral amino acids in both the presence and absence of Na+ and by the effect of N-ethylmaleimide. Transport data correlated to expression of genes encoding for CAT-1, CAT-2, CD-98, and y+LAT-2. A similar expression profile was detected in rat adrenal zona fasciculata. In addition, cationic amino acid uptake in Y1 cells was upregulated by ACTH and/or cAMP with a concomitant increase in nitric oxide (NO) production.

scholarly journals Sialic acid acquisition in bacteria–one substrate, many transporters

2016 ◽  
Vol 44 (3) ◽  
pp. 760-765 ◽  
Author(s):  
Gavin H. Thomas
Keyword(s):  
Sialic Acid ◽  
Immune Evasion ◽  
Sialic Acids ◽  
Major Facilitator Superfamily ◽  
Acid Uptake ◽  
Gram Negative Bacteria ◽  
Acetylneuraminic Acid ◽  
Wide Range ◽  
Major Facilitator

The sialic acids are a family of 9-carbon sugar acids found predominantly on the cell-surface glycans of humans and other animals within the Deuterostomes and are also used in the biology of a wide range of bacteria that often live in association with these animals. For many bacteria sialic acids are simply a convenient source of food, whereas for some pathogens they are also used in immune evasion strategies. Many bacteria that use sialic acids derive them from the environment and so are dependent on sialic acid uptake. In this mini-review I will describe the discovery and characterization of bacterial sialic acids transporters, revealing that they have evolved multiple times across multiple diverse families of transporters, including the ATP-binding cassette (ABC), tripartite ATP-independent periplasmic (TRAP), major facilitator superfamily (MFS) and sodium solute symporter (SSS) transporter families. In addition there is evidence for protein-mediated transport of sialic acids across the outer membrane of Gram negative bacteria, which can be coupled to periplasmic processing of different sialic acids to the most common form, β-D-N-acetylneuraminic acid (Neu5Ac) that is most frequently taken up into the cell.

scholarly journals YjhS (NanS) Is Required for Escherichia coli To Grow on 9-O-Acetylated N-Acetylneuraminic Acid

2009 ◽  
Vol 191 (22) ◽  
pp. 7134-7139 ◽  
Author(s):  
Susan M. Steenbergen ◽  
Jamie L. Jirik ◽  
Eric R. Vimr
Keyword(s):  
Escherichia Coli ◽  
Sialic Acid ◽  
Mammalian Host ◽  
E Coli ◽  
Acetylneuraminic Acid

ABSTRACT The nanATEK-yhcH, yjhATS, and yjhBC operons in Escherichia coli are coregulated by environmental N-acetylneuraminic acid, the most prevalent sialic acid in nature. Here we show that YjhS (NanS) is a probable 9-O-acetyl N-acetylneuraminic acid esterase required for E. coli to grow on this alternative sialic acid, which is commonly found in mammalian host mucosal sites.

N-Acetylneuraminic acid uptake inPasteurella (Mannheimia) haemolyticaA2 occurs by an inducible and specific transport system

FEBS Letters ◽  
2001 ◽  
Vol 509 (1) ◽  
pp. 41-46 ◽  
Author(s):  
Sonia Solana ◽  
Ángel Reglero ◽  
Honorina Martı́nez-Blanco ◽  
Beatriz Revilla-Nuin ◽  
Ignacio G Bravo ◽  
...  
Keyword(s):  
Transport System ◽  
Acid Uptake ◽  
Acetylneuraminic Acid ◽  
Specific Transport ◽  
Specific Transport System

scholarly journals Identification and Biochemical Characterization of the Novel α2,3-Sialyltransferase WbwA from Pathogenic Escherichia coli Serotype O104

2015 ◽  
Vol 197 (24) ◽  
pp. 3760-3768 ◽  
Author(s):  
Diana Czuchry ◽  
Paul Desormeaux ◽  
Melissa Stuart ◽  
Donald L. Jarvis ◽  
Khushi L. Matta ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Sialic Acid ◽  
Biochemical Characterization ◽  
Enzyme Reaction ◽  
T Antigen ◽  
Glycan Structure ◽  
Content Type ◽  
E Coli ◽  
Pathogenic Escherichia Coli

ABSTRACTThe sialyl-T antigen sialylα2-3Galβ1-3GalNAc is a common O-glycan structure in human glycoproteins and is synthesized by sialyltransferase ST3Gal1. The enterohemorrhagicEscherichia coliserotype O104 has the rare ability to synthesize a sialyl-T antigen mimic. We showed here that thewbwAgene of theE. coliO104 antigen synthesis gene cluster encodes an α2,3-sialyltransferase WbwA that transfers sialic acid from CMP-sialic acid to Galβ1-3GalNAcα-diphosphate-lipid acceptor. Nuclear magnetic resonance (NMR) analysis of purified WbwA enzyme reaction product indicated that the sialyl-T antigen sialylα2-3Galβ1-3GalNAcα-diphosphate-lipid was synthesized. We showed that the conserved His-Pro (HP) motif and Glu/Asp residues of two EDG motifs in WbwA are important for the activity. The characterization studies showed that WbwA fromE. coliO104 is a monofunctional α2,3-sialyltransferase and is distinct from human ST3Gal1 as well as all other known sialyltransferases due to its unique acceptor specificity. This work contributes to knowledge of the biosynthesis of bacterial virulence factors.IMPORTANCEThis is the first characterization of a sialyltransferase involved in the synthesis of an O antigen inE. coli. The enzyme contributes to the mimicry of human sialyl-T antigen and has unique substrate specificity but very little sequence identity to other sialyltransferases. Thus, the bacterial sialyltransferase is related to the human counterpart only by the similarity of biochemical activity.

scholarly journals THE LYSOSOME PERIPHERY: BIOCHEMICAL AND ELECTROKINETIC PROPERTIES OF THE TRITOSOME SURFACE

1974 ◽  
Vol 60 (3) ◽  
pp. 764-773 ◽  
Author(s):  
Douglas M. Gersten ◽  
Thomas W. Kimmerer ◽  
H. Bruce Bosmann
Keyword(s):  
Sialic Acid ◽  
Electrophoretic Mobility ◽  
Membrane Fraction ◽  
Osmotic Shock ◽  
Particle Surface ◽  
Gas Liquid Chromatography ◽  
Present System ◽  
Acid Hydrolases ◽  
Acetylneuraminic Acid ◽  
Total Sialic Acid

Normal rat liver lysosomes were isolated by the technique of loading with Triton WR-1339. Purity of the preparation was monitored with marker enzymes; a high enrichment in acid hydrolases was obtained in the tritosome fraction. In 0.0145 M NaCl, 4.5% sorbitol, 0.6 mM NaHCO3, pH 7.2 at 25°C the tritosomes had an electrophoretic mobility of -1.77 ± 0.02 µm/s/V/cm, a zeta potential of 23.2 mV, a surface charge of 1970 esu/cm2, and 33,000 electrons per particle surface assuming a tritosome diameter of 5 x 10-7 m. Treatment of the tritosomes with 50 µg neuraminidase/mg tritosome protein lowered the electrophoretic mobility of the tritosome to -1.23 ± 0.02 µm/s/V/cm under the same conditions and caused the release of 2.01 µg sialic acid/mg tritosome protein. Treatment of the tritosomes with hyaluronidase did not affect their electrophoretic mobility, while trypsin treatment elevated the net negative electrophoretic mobility of the tritosomes. Tritosome electrophoretic mobilities indicated a homogeneous tritosome population and varied greatly with ionic strength of the suspending media. pH vs. electrophoretic mobility curves indicated the tritosome periphery to contain an acid-dissociable group which likely represents the carboxyl group of N-acetylneuraminic acid; this was not conclusively proven, however, since the tritosomes lysed below a pH of 4 in the present system. Total tritosome carbohydrate (anthrone-positive material as glucose equivalents) was 0.19 mg/mg tritosome protein while total sialic acid was 3.8 µg (11.4 nmol)/mg tritosome protein. A tritosome "membrane" fraction was prepared by osmotic shock, homogenization, and sedimentation. Approximately 25% of the total tritosome protein was present in this fraction. Analysis by gas-liquid chromatography and amino acid analyzer showed the following carbohydrate composition of the tritosome membrane fraction (in microgram per milligram tritosome membrane protein): N-acetylneuraminic acid, 14.8 ± 3; glucosamine, 24 ± 3; galactosamine, 10 ± 2; glucose, 21 ± 2; galactose, 26 ± 2; mannose, 31 ± 5; fucose, 7 ± 1; xylose, 0; and arabinose, 0. The results indicate that the tritosome periphery is characterized by external terminal sialic acid residues and an extensive complement of glycoconjugates. Essentially all the tritosome N-acetylneuraminic acid is located in the membrane and about 53% of it is neuraminidase susceptible.

scholarly journals Artificially induced active transport of amino acid driven by the efflux of a sugar via a heterologous transport system in de-energized Escherichia coli

1979 ◽  
Vol 178 (1) ◽  
pp. 103-107 ◽  
Author(s):  
M Bentaboulet ◽  
A Robin ◽  
A Kepes
Keyword(s):  
Escherichia Coli ◽  
Amino Acid ◽  
Active Transport ◽  
Transport System ◽  
Coupling Efficiency ◽  
Amino Acid Uptake ◽  
Energy Sources ◽  
Acid Uptake ◽  
Complex Sequence ◽  
The Poor

Consistent with the model of an H+ cotransport, amino acid uptake can be driven by a proton gradient generated by an efflux of sugar when the normal energy sources are suppressed. Heterologous countertransport is completely inhibited by uncouplers unlike homologous countertransport. Positive coupling was obtained with methyl thiogalactoside/proline, methyl thiogalactoside/phenylalanine, gluconate/proline; however, the poor coupling efficiency suggests a more complex sequence of reactions.

scholarly journals Succinate uptake and related proton movements in Escherichia coli K12

1975 ◽  
Vol 152 (3) ◽  
pp. 647-654 ◽  
Author(s):  
S J Gutowski ◽  
H Rosenberg
Keyword(s):  
Escherichia Coli ◽  
Dicarboxylic Acid ◽  
Transport System ◽  
Dicarboxylic Acids ◽  
Acid Transport ◽  
Cationic Form ◽  
Whole Cells ◽  
Escherichia Coli K12 ◽  
Proton Uptake

1. The apparent Km values for succinate uptake by whole cells of Escherichia coli K12 depend on pH in the range 6.5-7.4.2. Uptake of succinate in lightly buffered medium is accompanied by proton uptake. 3. The apparent Km values for succinate uptake and for succinate-induced proton uptake are similar. 4. Approximately two protons enter the cell with each succinate molecule. 5. The pattern of inhibition of succinate uptake is similar to that of succinate-induced proton uptake. 6. Uptake of fumarate and malate, which share the succinate-transport system, is also accompanied by the uptake of approximately two protons per molecule of fumarate or malate. 7. Uptake of aspartate by the dicarboxylic acid-transport system is accompanied by the uptake of approximatley two protons per molecule of asparatate. 8. It is concluded that uptake of dicarboxylic acids by the dicarboxylic acid-transport system is obligatorily coupled to proton uptake such that succinate, malate and fumarate are taken up in electroneutral form and asparate is taken up in cationic form. 9. These results are consistent with, though they do not definitely prove, the energization of succinate uptake of the deltapH.

scholarly journals Novel Sialic Acid Transporter of Haemophilus influenzae

2005 ◽  
Vol 73 (9) ◽  
pp. 5291-5300 ◽  
Author(s):  
Simon Allen ◽  
Anthony Zaleski ◽  
Jason W. Johnston ◽  
Bradford W. Gibson ◽  
Michael A. Apicella
Keyword(s):  
Sialic Acid ◽  
Haemophilus Influenzae ◽  
Opportunistic Pathogen ◽  
Chronic Obstructive ◽  
Acid Uptake ◽  
Biofilm Growth ◽  
Obstructive Pulmonary Disease ◽  
Acetylneuraminic Acid ◽  
Acid Incorporation ◽  
Transporter Family

ABSTRACT Nontypeable Haemophilus influenzae is an opportunistic pathogen and a common cause of otitis media in children and of chronic bronchitis and pneumonia in patients with chronic obstructive pulmonary disease. The lipooligosaccharides, a major component of the outer membrane of H. influenzae, play an important role in microbial virulence and pathogenicity. N-Acetylneuraminic acid (sialic acid) can be incorporated into the lipooligosaccharides as a terminal nonreducing sugar. Although much of the pathway of sialic acid incorporation into lipooligosaccharides is understood, the transporter responsible for N-acetylneuraminic acid uptake in H. influenzae has yet to be characterized. In this paper we demonstrate that this transporter is a novel sugar transporter of the tripartite ATP-independent periplasmic transporter family. In the absence of this transporter, H. influenzae cannot incorporate sialic acid into its lipooligosaccharides, making the organism unable to survive when exposed to human serum and causing reduced viability in biofilm growth.

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