scholarly journals The Essential tacF Gene Is Responsible for the Choline-Dependent Growth Phenotype of Streptococcus pneumoniae

2007 ◽  
Vol 189 (19) ◽  
pp. 7105-7111 ◽  
Author(s):  
Marlen Damjanovic ◽  
Arun S. Kharat ◽  
Alice Eberhardt ◽  
Alexander Tomasz ◽  
Waldemar Vollmer
Keyword(s):  
Cell Wall ◽  
Streptococcus Pneumoniae ◽  
Teichoic Acid ◽  
Laboratory Strain ◽  
Nutritional Requirement ◽  
Pcr Product ◽  
Dependent Growth ◽  
Strict Specificity ◽  
Transmembrane Transporters ◽  
Independent Mutant

ABSTRACT Streptococcus pneumoniae has an absolute nutritional requirement for choline, and the choline molecules are known to incorporate exclusively into the cell wall and membrane teichoic acids of the bacterium. We describe here the isolation of a mutant of strain R6 in which a single G→T point mutation in the gene tacF (formerly designated spr1150) is responsible for generating a choline-independent phenotype. The choline-independent phenotype could be transferred to the laboratory strain R6 and to the encapsulated strain D39 by genetic transformation with a PCR product or with a plasmid carrying the mutated tacF gene. The tacF gene product belongs to the protein family of polysaccharide transmembrane transporters (flippases). A model is presented in which TacF is required for the transport of the teichoic acid subunits across the cytoplasmic membrane. According to this model, wild-type TacF has a strict specificity for choline-containing subunits, whereas the TacF present in the choline-independent mutant strain is able to transport both choline-containing and choline-free teichoic acid chains. The proposed transport specificity of parental-type TacF for choline-containing subunits would ensure the loading of the cell wall with teichoic acid chains decorated with choline residues, which appear to be essential for the virulence of this pathogen.

scholarly journals Mutations in the tacF Gene of Clinical Strains and Laboratory Transformants of Streptococcus pneumoniae: Impact on Choline Auxotrophy and Growth Rate

2008 ◽  
Vol 190 (12) ◽  
pp. 4129-4138 ◽  
Author(s):  
Ana González ◽  
Daniel Llull ◽  
María Morales ◽  
Pedro García ◽  
Ernesto García
Keyword(s):  
Streptococcus Pneumoniae ◽  
Repeat Unit ◽  
Teichoic Acid ◽  
Integral Membrane Protein ◽  
Nutritional Requirement ◽  
Spontaneous Mutant ◽  
Free Medium ◽  
Streptococcus Oralis ◽  
Complex Relationships ◽  
Lipoteichoic Acids

ABSTRACT The nutritional requirement that Streptococcus pneumoniae has for the aminoalcohol choline as a component of teichoic and lipoteichoic acids appears to be exclusive to this prokaryote. A mutation in the tacF gene, which putatively encodes an integral membrane protein (possibly, a teichoic acid repeat unit transporter), has been recently identified as responsible for generating a choline-independent phenotype of S. pneumoniae (M. Damjanovic, A. S. Kharat, A. Eberhardt, A. Tomasz, and W. Vollmer, J. Bacteriol. 189:7105-7111, 2007). We now report that Streptococcus mitis can grow in choline-free medium, as previously illustrated for Streptococcus oralis. While we confirmed the finding by Damjanovic et al. of the involvement of TacF in the choline dependence of the pneumococcus, the genetic transformation of S. pneumoniae R6 by using S. mitis SK598 DNA and several PCR-amplified tacF fragments suggested that a minimum of two mutations were required to confer improved fitness to choline-independent S. pneumoniae mutants. This conclusion is supported by sequencing results also reported here that indicate that a spontaneous mutant of S. pneumoniae (strain JY2190) able to proliferate in the absence of choline (or analogs) is also a double mutant for the tacF gene. Microscopic observations and competition experiments during the cocultivation of choline-independent strains confirmed that a minimum of two amino acid changes were required to confer improved fitness to choline-independent pneumococcal strains when growing in medium lacking any aminoalcohol. Our results suggest complex relationships among the different regions of the TacF teichoic acid repeat unit transporter.

Specific and spatial labeling of choline-containing teichoic acids in Streptococcus pneumoniae by click chemistry

2017 ◽  
Vol 53 (76) ◽  
pp. 10572-10575 ◽  
Author(s):  
A. M. Di Guilmi ◽  
J. Bonnet ◽  
S. Peiβert ◽  
C. Durmort ◽  
B. Gallet ◽  
...  
Keyword(s):  
Cell Wall ◽  
Streptococcus Pneumoniae ◽  
Click Chemistry ◽  
Metabolic Pathway ◽  
Teichoic Acid ◽  
Teichoic Acids ◽  
First Time

A choline metabolic pathway was exploited to label for the first time teichoic acid (TA) in the Streptococcus pneumoniae cell wall.

Differential Release of Lipoteichoic and Teichoic Acids from Streptococcus pneumoniae as a Result of Exposure to β-Lactam Antibiotics, Rifamycins, Trovafloxacin, and Quinupristin-Dalfopristin

1998 ◽  
Vol 42 (2) ◽  
pp. 277-281 ◽  
Author(s):  
K. Stuertz ◽  
H. Schmidt ◽  
H. Eiffert ◽  
P. Schwartz ◽  
M. Mäder ◽  
...  
Keyword(s):  
Cell Wall ◽  
Streptococcus Pneumoniae ◽  
Antibacterial Agents ◽  
Teichoic Acid ◽  
Lipoteichoic Acid ◽  
Enzyme Linked Immunosorbent Assay ◽  
Low Concentrations ◽  
Type 3 ◽  
Antibiotic Dose ◽  
Wall Components

ABSTRACT The release of lipoteichoic acid (LTA) and teichoic acid (TA) from a Streptococcus pneumoniae type 3 strain during exposure to ceftriaxone, meropenem, rifampin, rifabutin, quinupristin-dalfopristin, and trovafloxacin in tryptic soy broth was monitored by a newly developed enzyme-linked immunosorbent assay. At a concentration of 10 μg/ml, a rapid and intense release of LTA and TA occurred during exposure to ceftriaxone (3,248 ± 1,688 ng/ml at 3 h and 3,827 ± 2,133 ng/ml at 12 h) and meropenem (2,464 ± 1,081 ng/ml at 3 h and 2,900 ± 1,364 ng/ml at 12 h). Three hours after exposure to rifampin, rifabutin, quinupristin-dalfopristin, and trovafloxacin, mean LTA and TA concentrations of less than 460 ng/ml were observed (for each group,P < 0.01 versus the concentrations after exposure to ceftriaxone). After 12 h of treatment, the LTA and TA concentrations were 463 ± 126 ng/ml after exposure to rifampin, 669 ± 303 ng/ml after exposure to rifabutin, and 1,236 ± 772 ng/ml after exposure to quinupristin-dalfopristin (for each group,P < 0.05 versus the concentrations after exposure to ceftriaxone) and 1,745 ± 1,185 ng/ml after exposure to trovafloxacin (P = 0.12 versus the concentration after exposure to ceftriaxone). At 10 μg/ml, bactericidal antibacterial agents that do not primarily affect cell wall synthesis reduced the amount of LTA and TA released during their cidal action againstS. pneumoniae in comparison with the amount released after exposure to β-lactams. Larger quantities of LTA and TA were released after treatment with low concentrations (1× the MIC and 1× the minimum bactericidal concentration) than after no treatment for all antibacterial agents except the rifamycins. This does not support the concept of using a low first antibiotic dose to prevent the release of proinflammatory cell wall components.

DiiA is a novel dimorphic cell wall protein of Streptococcus pneumoniae involved in invasive disease

2016 ◽  
Vol 73 (1) ◽  
pp. 71-81 ◽  
Author(s):  
María S. Escolano-Martínez ◽  
Arnau Domenech ◽  
José Yuste ◽  
María I. Cercenado ◽  
Carmen Ardanuy ◽  
...  
Keyword(s):  
Cell Wall ◽  
Streptococcus Pneumoniae ◽  
Invasive Disease ◽  
Cell Wall Protein

scholarly journals Structural and Functional Insights into Peptidoglycan Access for the Lytic Amidase LytA of Streptococcus pneumoniae

mBio ◽  
2014 ◽  
Vol 5 (1) ◽  
Author(s):  
Peter Mellroth ◽  
Tatyana Sandalova ◽  
Alexey Kikhney ◽  
Francisco Vilaplana ◽  
Dusan Hesek ◽  
...  
Keyword(s):  
Cell Wall ◽  
Streptococcus Pneumoniae ◽  
Solution Structure ◽  
Substrate Binding ◽  
Substrate Recognition ◽  
Lytic Activity ◽  
Site Directed Mutagenesis ◽  
Binding Motif ◽  
Content Type ◽  
Peptidoglycan Synthesis

ABSTRACT The cytosolic N-acetylmuramoyl-l-alanine amidase LytA protein of Streptococcus pneumoniae, which is released by bacterial lysis, associates with the cell wall via its choline-binding motif. During exponential growth, LytA accesses its peptidoglycan substrate to cause lysis only when nascent peptidoglycan synthesis is stalled by nutrient starvation or β-lactam antibiotics. Here we present three-dimensional structures of LytA and establish the requirements for substrate binding and catalytic activity. The solution structure of the full-length LytA dimer reveals a peculiar fold, with the choline-binding domains forming a rigid V-shaped scaffold and the relatively more flexible amidase domains attached in a trans position. The 1.05-Å crystal structure of the amidase domain reveals a prominent Y-shaped binding crevice composed of three contiguous subregions, with a zinc-containing active site localized at the bottom of the branch point. Site-directed mutagenesis was employed to identify catalytic residues and to investigate the relative impact of potential substrate-interacting residues lining the binding crevice for the lytic activity of LytA. In vitro activity assays using defined muropeptide substrates reveal that LytA utilizes a large substrate recognition interface and requires large muropeptide substrates with several connected saccharides that interact with all subregions of the binding crevice for catalysis. We hypothesize that the substrate requirements restrict LytA to the sites on the cell wall where nascent peptidoglycan synthesis occurs. IMPORTANCE Streptococcus pneumoniae is a human respiratory tract pathogen responsible for millions of deaths annually. Its major pneumococcal autolysin, LytA, is required for autolysis and fratricidal lysis and functions as a virulence factor that facilitates the spread of toxins and factors involved in immune evasion. LytA is also activated by penicillin and vancomycin and is responsible for the lysis induced by these antibiotics. The factors that regulate the lytic activity of LytA are unclear, but it was recently demonstrated that control is at the level of substrate recognition and that LytA required access to the nascent peptidoglycan. The present study was undertaken to structurally and functionally investigate LytA and its substrate-interacting interface and to determine the requirements for substrate recognition and catalysis. Our results reveal that the amidase domain comprises a complex substrate-binding crevice and needs to interact with a large-motif epitope of peptidoglycan for catalysis.

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