scholarly journals The d-Xylose-Binding Protein, XylF, from Thermoanaerobacter ethanolicus 39E: Cloning, Molecular Analysis, and Expression of the Structural Gene

1998 ◽  
Vol 180 (14) ◽  
pp. 3570-3577 ◽  
Author(s):  
Milutin Erbeznik ◽  
Herbert J. Strobel ◽  
Karl A. Dawson ◽  
Chris R. Jones
Keyword(s):  
Amino Acid ◽  
Binding Proteins ◽  
Binding Protein ◽  
Xylose Isomerase ◽  
Thermophilic Bacterium ◽  
Transport Systems ◽  
Uptake System ◽  
Calculated Molecular Mass ◽  
Thermoanaerobacter Ethanolicus ◽  
High Affinity

ABSTRACT Immediately downstream from the Thermoanaerobacter ethanolicus xylAB operon, comprising genes that encoded-xylose isomerase and d-xylulose kinase, lies a 1,101-bp open reading frame that exhibits 61% amino acid sequence identity to the Escherichia coli d-xylose binding periplasmic receptor, XylF, a component of the high-affinity binding-protein-dependent d-xylose transport. The 25-residue N-terminal fragment of the deduced T. ethanolicus XylF has typical features of bacterial leader peptides. The C-terminal portion of this leader sequence matches the cleavage consensus for lipoproteins and is followed by a 22-residue putative linker sequence rich in serine, threonine, and asparagine. The putative mature 341-amino-acid-residue XylF (calculated molecular mass of 37,069 Da) appears to be a lipoprotein attached to the cell membrane via a lipid anchor covalently linked to the N-terminal cysteine, as demonstrated by metabolic labelling of the recombinant XylF with [14C]palmitate. The induced E. coli avidly bound d-[14C]xylose, yielding additional evidence that T. ethanolicus XylF is thed-xylose-binding protein. On the basis of sequence comparison of XylFs to other monosaccharide-binding proteins, we propose that the sequence signature of binding proteins specific for hexoses and pentoses be refined as (KDQ)(LIVFAG)3IX3(DN)(SGP)X3(GS)X(LIVA)2X2A. Transcription of the monocistronic 1.3-kb xylF mRNA is inducible by xylose and unaffected by glucose. Primer extension analysis indicated that xylF transcription initiates from two +1 sites, both situated within the xylAB operon. Unlike in similar transport systems in other bacteria, the genes specifying the membrane components (e.g., ATP-binding protein and permease) of the high-affinity d-xylose uptake system are not located in the vicinity of xylF in T. ethanolicus. This is the first report of a gene encoding a xylose-binding protein in a gram-positive or thermophilic bacterium.

scholarly journals Increasing Ceftriaxone Resistance and Multiple Alterations of Penicillin-Binding Proteins among Penicillin-Resistant Streptococcus pneumoniae Isolates in Taiwan

2007 ◽  
Vol 51 (9) ◽  
pp. 3404-3406 ◽  
Author(s):  
Cheng-Hsun Chiu ◽  
Lin-Hui Su ◽  
Yhu-Chering Huang ◽  
Jui-Chia Lai ◽  
Hsiu-Ling Chen ◽  
...  
Keyword(s):  
Streptococcus Pneumoniae ◽  
Amino Acid ◽  
Binding Proteins ◽  
Binding Protein ◽  
Penicillin Binding Protein ◽  
Penicillin Binding Proteins

ABSTRACT The rate of nonsusceptibility of penicillin-resistant Streptococcus pneumoniae strains to ceftriaxone increased significantly in Taiwan in 2005. Approximately 90% of the ceftriaxone-nonsusceptible isolates were found to be of four major serotypes (serotypes 6B, 14, 19F, and 23F). Seven amino acid alterations in the penicillin-binding protein 2B transpeptidase-encoding region specifically contributed to the resistance.

Localization of the high-affinity glutamate transporter EAAC1 in rat kidney

1997 ◽  
Vol 273 (6) ◽  
pp. F1023-F1029 ◽  
Author(s):  
Chairat Shayakul ◽  
Yoshikatsu Kanai ◽  
Wen-Sen Lee ◽  
Dennis Brown ◽  
Jeffrey D. Rothstein ◽  
...  
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Rat Kidney ◽  
Cell Volume Regulation ◽  
Glutamate Transporter ◽  
Transport Systems ◽  
Kidney Cortex ◽  
Acid Base Balance ◽  
Western Blots ◽  
High Affinity

Most amino acids filtered by the glomerulus are reabsorbed in the kidney via specialized transport systems. Recently, the cDNA encoding a high-affinity glutamate transporter, EAAC1, has been isolated and shown to be expressed at high levels in the kidney. To determine the potential role of EAAC1 in renal acidic amino acid reabsorption, the distribution of EAAC1 mRNA and protein in rat kidney was examined. In situ hybridization revealed that EAAC1 mRNA is expressed predominantly in S2 and S3 segments of the proximal tubules and at low levels in the inner stripe of outer medulla and inner medulla. Polyclonal antibodies raised against the carboxy terminus of EAAC1 recognized a single band of ∼70 kDa on Western blots of membrane protein from kidney cortex and medulla. Immunofluorescence microscopy revealed intense signals in the luminal membrane of S2 and S3 segments and weaker signals in S1 segments, descending thin limbs of long-loop nephrons, medullary thick ascending limbs, and distal convoluted tubules. These results are consistent with EAAC1 encoding the previously described apical high-affinity glutamate transporter in the kidney that mediates reabsorption of acidic amino acids in tubules beyond early proximal tubule S1 segments. Potential additional roles of EAAC1 in acid/base balance, cell volume regulation, and amino acid metabolism are discussed.

scholarly journals Znu Is the Predominant Zinc Importer in Yersinia pestis during In Vitro Growth but Is Not Essential for Virulence

2010 ◽  
Vol 78 (12) ◽  
pp. 5163-5177 ◽  
Author(s):  
Daniel C. Desrosiers ◽  
Scott W. Bearden ◽  
Ildefonso Mier ◽  
Jennifer Abney ◽  
James T. Paulley ◽  
...  
Keyword(s):  
Yersinia Pestis ◽  
Binding Protein ◽  
Bacterial Pathogens ◽  
Titration Calorimetry ◽  
Growth Defect ◽  
Uptake System ◽  
High Affinity ◽  
Zn Uptake ◽  
Low Concentrations

ABSTRACT Little is known about Zn homeostasis in Yersinia pestis, the plague bacillus. The Znu ABC transporter is essential for zinc (Zn) uptake and virulence in a number of bacterial pathogens. Bioinformatics analysis identified ZnuABC as the only apparent high-affinity Zn uptake system in Y. pestis. Mutation of znuACB caused a growth defect in Chelex-100-treated PMH2 growth medium, which was alleviated by supplementation with submicromolar concentrations of Zn. Use of transcriptional reporters confirmed that Zur mediated Zn-dependent repression and that it can repress gene expression in response to Zn even in the absence of Znu. Virulence testing in mouse models of bubonic and pneumonic plague found only a modest increase in survival in low-dose infections by the znuACB mutant. Previous studies of cluster 9 (C9) transporters suggested that Yfe, a well-characterized C9 importer for manganese (Mn) and iron in Y. pestis, might function as a second, high-affinity Zn uptake system. Isothermal titration calorimetry revealed that YfeA, the solute-binding protein component of Yfe, binds Mn and Zn with comparably high affinities (dissociation constants of 17.8 ± 4.4 nM and 6.6 ± 1.2 nM, respectively), although the complete Yfe transporter could not compensate for the loss of Znu in in vitro growth studies. Unexpectedly, overexpression of Yfe interfered with the znu mutant's ability to grow in low concentrations of Zn, while excess Zn interfered with the ability of Yfe to import iron at low concentrations; these results suggest that YfeA can bind Zn in the bacterial cell but that Yfe is incompetent for transport of the metal. In addition to Yfe, we have now eliminated MntH, FetMP, Efe, Feo, a substrate-binding protein, and a putative nickel transporter as the unidentified, secondary Zn transporter in Y. pestis. Unlike other bacterial pathogens, Y. pestis does not require Znu for high-level infectivity and virulence; instead, it appears to possess a novel class of transporter, which can satisfy the bacterium's Zn requirements under in vivo metal-limiting conditions. Our studies also underscore the need for bacterial cells to balance binding and transporter specificities within the periplasm in order to maintain transition metal homeostasis.

scholarly journals Affinity of Tomopenem (CS-023) for Penicillin-Binding Proteins in Staphylococcus aureus, Escherichia coli, and Pseudomonas aeruginosa

2008 ◽  
Vol 53 (3) ◽  
pp. 1238-1241 ◽  
Author(s):  
Tetsufumi Koga ◽  
Chika Sugihara ◽  
Masayo Kakuta ◽  
Nobuhisa Masuda ◽  
Eiko Namba ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Staphylococcus Aureus ◽  
Pseudomonas Aeruginosa ◽  
Binding Proteins ◽  
Binding Protein ◽  
Penicillin Binding Protein ◽  
Penicillin Binding Proteins ◽  
E Coli ◽  
High Affinity

ABSTRACT Tomopenem (formerly CS-023), a novel 1β-methylcarbapenem, exhibited high affinity for penicillin-binding protein (PBP) 2 in Staphylococcus aureus, PBP 2 in Escherichia coli, and PBPs 2 and 3 in Pseudomonas aeruginosa, which are considered major lethal targets. Morphologically, tomopenem induced spherical forms in E. coli and short filamentation with bulges in P. aeruginosa, which correlated with the drug's PBP profiles. The potential of resistance of these bacteria to tomopenem was comparable to that to imipenem.

scholarly journals Genetic and Biochemical Characterization of a High-Affinity Betaine Uptake System (BusA) in Lactococcus lactisReveals a New Functional Organization within Bacterial ABC Transporters

1999 ◽  
Vol 181 (20) ◽  
pp. 6238-6246 ◽  
Author(s):  
David Obis ◽  
Alain Guillot ◽  
Jean-Claude Gripon ◽  
Pierre Renault ◽  
Alexander Bolotin ◽  
...  
Keyword(s):  
Abc Transporters ◽  
Biochemical Characterization ◽  
Transmembrane Domain ◽  
Functional Organization ◽  
Binding Protein ◽  
Atp Binding ◽  
Uptake System ◽  
High Affinity ◽  
Atp Binding Protein

ABSTRACT The cytoplasmic accumulation of exogenous betaine stimulates the growth of Lactococcus lactis cultivated under hyperosmotic conditions. We report that L. lactis possesses a single betaine transport system that belongs to the ATP-binding cassette (ABC) superfamily of transporters. Through transposon mutagenesis, a mutant deficient in betaine transport was isolated. We identified two genes, busAA and busAB, grouped in an operon, busA (betaine uptake system). The transcription of busA is strongly regulated by the external osmolality of the medium. The busAA gene codes for the ATP-binding protein. busAB encodes a 573-residue polypeptide which presents two striking features: (i) a fusion between the regions encoding the transmembrane domain (TMD) and the substrate-binding domain (SBD) and (ii) a swapping of the SBD subdomains when compared to the Bacillus subtilisbetaine-binding protein, OpuAC. BusA of L. lactis displays a high affinity towards betaine (Km = 1.7 μM) and is an osmosensor whose activity is tightly regulated by external osmolality, leading the betaine uptake capacity ofL. lactis to be under dual control at the biochemical and genetic levels. A protein presenting the characteristics predicted for BusAB was detected in the membrane fraction of L. lactis. The fusion between the TMD and the SBD is the first example of a new organization within prokaryotic ABC transporters.

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