Document Detail


Characterization of a Pseudomonas putida ABC transporter (AatJMQP) required for acidic amino acid uptake: biochemical properties and regulation by the Aau two-component system.
MedLine Citation:
PMID:  18310026     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
We describe an ATP-binding cassette (ABC) transporter in Pseudomonas putida KT2440 that mediates the uptake of glutamate and aspartate. The system (AatJMQP, for acidic amino acid transport) is encoded by an operon involving genes PP1071-PP1068. A deletion mutant with inactivated solute-binding protein (KTaatJ) failed to grow on Glu and Gln as sole sources of carbon and nitrogen, while a mutant lacking a functional nucleotide-binding domain (KTaatP) was able to adapt to growth on Glu after an extended lag phase. Uptake of Glu and Asp by either mutant was greatly impaired at both low and high amino acid concentrations. The purified solute-binding protein AatJ exhibited high affinity towards Glu and Asp (K(d)=0.4 and 1.3 muM, respectively), while Gln and Asn as well as dicarboxylates (succinate and fumarate) were bound with much lower affinity. We further show that the expression of AatJMQP is controlled by the sigma(54)-dependent two-component system AauRS. Binding of the response regulator AauR to the aat promoter was examined by gel mobility shift assays and DNase I footprinting. By in silico screening, the AauR-binding motif (the inverted repeat TTCGGNNNNCCGAA) was detected in further P. putida KT2440 genes with established or putative functions in acidic amino acid utilization, and also occurred in other pseudomonads. The products of these AauR-responsive genes include the H(+)/Glu symporter GltP, a periplasmic glutaminase/asparaginase, AnsB, and phosphoenolpyruvate synthase (PpsA), a key enzyme of gluconeogenesis in Gram-negative bacteria. Based on these findings, we propose that AauR is a central regulator of acidic amino acid uptake and metabolism in pseudomonads.
Authors:
Birendra Singh; Klaus-Heinrich Röhm
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't    
Journal Detail:
Title:  Microbiology (Reading, England)     Volume:  154     ISSN:  1350-0872     ISO Abbreviation:  Microbiology (Reading, Engl.)     Publication Date:  2008 Mar 
Date Detail:
Created Date:  2008-03-03     Completed Date:  2008-05-01     Revised Date:  2008-11-21    
Medline Journal Info:
Nlm Unique ID:  9430468     Medline TA:  Microbiology     Country:  England    
Other Details:
Languages:  eng     Pagination:  797-809     Citation Subset:  IM    
Affiliation:
Philipps University, Institute of Physiological Chemistry, D-35043 Marburg, Germany.
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MeSH Terms
Descriptor/Qualifier:
ATP-Binding Cassette Transporters / genetics,  metabolism*
Aspartic Acid / metabolism*
Bacterial Proteins / metabolism
Base Sequence
Binding Sites
DNA Footprinting
DNA, Bacterial / metabolism
DNA-Binding Proteins / metabolism*
Electrophoretic Mobility Shift Assay
Gene Deletion
Gene Expression Regulation, Bacterial / physiology*
Gene Order
Glutamic Acid / metabolism*
Models, Biological
Molecular Sequence Data
Operon
Promoter Regions, Genetic
Protein Binding
Pseudomonas putida / genetics,  growth & development,  metabolism*
Signal Transduction
Substrate Specificity
Chemical
Reg. No./Substance:
0/ATP-Binding Cassette Transporters; 0/Bacterial Proteins; 0/DNA, Bacterial; 0/DNA-Binding Proteins; 56-84-8/Aspartic Acid; 56-86-0/Glutamic Acid

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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