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000030967 0247_ $$2DOI$$a10.1021/bi0340595
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000030967 084__ $$2WoS$$aBiochemistry & Molecular Biology
000030967 1001_ $$0P:(DE-Juel1)VDB2886$$aGerharz, T.$$b0$$uFZJ
000030967 245__ $$aIdentification of basic amino acid residues important for citrate binding by the periplasmic receptor domain of the sensor kinase CitA
000030967 260__ $$aColumbus, Ohio$$bAmerican Chemical Society$$c2003
000030967 300__ $$a5917 - 5924
000030967 3367_ $$0PUB:(DE-HGF)16$$2PUB:(DE-HGF)$$aJournal Article
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000030967 440_0 $$0798$$aBiochemistry$$v42$$x0006-2960
000030967 500__ $$aRecord converted from VDB: 12.11.2012
000030967 520__ $$aThe sensor kinase CitA and the response regulator CitB of Klebsiella pneumoniae form the paradigm of a subfamily of bacterial two-component regulatory systems that are capable of sensing tri- or dicarboxylates in the environment and then induce transporters for the uptake of these compounds. We recently showed that the separated periplasmic domain of CitA, termed CitAP (encompasses residues 45-176 supplemented with an N-terminal methionine residue and a C-terminal hexahistidine tag), is a highly specific citrate receptor with a K(d) of 5.5 microM at pH 7. To identify positively charged residues involved in binding the citrate anion, each of the arginine, lysine, and histidine residues in CitAP was exchanged for alanine, and the resulting 17 muteins were analyzed by isothermal titration calorimetry (ITC). In 12 cases, the K(d) for citrate was identical to that of wild-type CitAP or slightly changed (3.9-17.2 microM). In one case (R98A), the K(d) was 6-fold decreased (0.8 microM), whereas in four cases (R66A, H69A, R107A, and K109A) the K(d) was 38- to >300-fold increased (0.2 to >1 mM). The secondary structure of the latter five proteins in their apo-form as deduced from far-UV circular dichroism (CD) spectra did not differ from the apo-form of wild-type CitAP; however, all of them showed an increased thermostability. Citrate increased the melting point (T(m)) of wild-type CitAP and mutein R98A by 6.2 and 9.5 degrees C, respectively, but had no effect on the T(m) of the four proteins with disturbed binding. Three of the residues important for citrate binding (R66, H69, and R107) are highly conserved in the CitA subfamily of sensor kinases, indicating that they might be involved in ligand binding by many of these sensor kinases.
000030967 536__ $$0G:(DE-Juel1)FUEK256$$2G:(DE-HGF)$$aBiotechnologie$$cL02$$x0
000030967 588__ $$aDataset connected to Web of Science, Pubmed
000030967 650_2 $$2MeSH$$aAmino Acid Sequence
000030967 650_2 $$2MeSH$$aAmino Acid Substitution
000030967 650_2 $$2MeSH$$aAmino Acids, Basic: chemistry
000030967 650_2 $$2MeSH$$aBacterial Proteins: chemistry
000030967 650_2 $$2MeSH$$aBacterial Proteins: genetics
000030967 650_2 $$2MeSH$$aBacterial Proteins: metabolism
000030967 650_2 $$2MeSH$$aBase Sequence
000030967 650_2 $$2MeSH$$aBinding Sites
000030967 650_2 $$2MeSH$$aCircular Dichroism
000030967 650_2 $$2MeSH$$aCitric Acid: metabolism
000030967 650_2 $$2MeSH$$aDNA, Bacterial: genetics
000030967 650_2 $$2MeSH$$aEscherichia coli Proteins: chemistry
000030967 650_2 $$2MeSH$$aEscherichia coli Proteins: genetics
000030967 650_2 $$2MeSH$$aEscherichia coli Proteins: metabolism
000030967 650_2 $$2MeSH$$aKinetics
000030967 650_2 $$2MeSH$$aKlebsiella pneumoniae: genetics
000030967 650_2 $$2MeSH$$aKlebsiella pneumoniae: metabolism
000030967 650_2 $$2MeSH$$aLigands
000030967 650_2 $$2MeSH$$aMolecular Sequence Data
000030967 650_2 $$2MeSH$$aMutagenesis, Site-Directed
000030967 650_2 $$2MeSH$$aPeriplasm: metabolism
000030967 650_2 $$2MeSH$$aProtein Binding
000030967 650_2 $$2MeSH$$aProtein Kinases: chemistry
000030967 650_2 $$2MeSH$$aProtein Kinases: genetics
000030967 650_2 $$2MeSH$$aProtein Kinases: metabolism
000030967 650_2 $$2MeSH$$aProtein Structure, Tertiary
000030967 650_2 $$2MeSH$$aSequence Homology, Amino Acid
000030967 650_2 $$2MeSH$$aSpectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
000030967 650_2 $$2MeSH$$aThermodynamics
000030967 650_2 $$2MeSH$$aTranscription Factors: chemistry
000030967 650_2 $$2MeSH$$aTranscription Factors: genetics
000030967 650_2 $$2MeSH$$aTranscription Factors: metabolism
000030967 650_7 $$00$$2NLM Chemicals$$aAmino Acids, Basic
000030967 650_7 $$00$$2NLM Chemicals$$aBacterial Proteins
000030967 650_7 $$00$$2NLM Chemicals$$aCitB protein, Klebsiella pneumoniae
000030967 650_7 $$00$$2NLM Chemicals$$aDNA, Bacterial
000030967 650_7 $$00$$2NLM Chemicals$$aEscherichia coli Proteins
000030967 650_7 $$00$$2NLM Chemicals$$aLigands
000030967 650_7 $$00$$2NLM Chemicals$$aTranscription Factors
000030967 650_7 $$077-92-9$$2NLM Chemicals$$aCitric Acid
000030967 650_7 $$0EC 2.7.-$$2NLM Chemicals$$aProtein Kinases
000030967 650_7 $$0EC 2.7.1.-$$2NLM Chemicals$$adpiB protein, E coli
000030967 650_7 $$2WoSType$$aJ
000030967 7001_ $$0P:(DE-HGF)0$$aReinelt, S.$$b1
000030967 7001_ $$0P:(DE-Juel1)VDB723$$aKaspar, S.$$b2$$uFZJ
000030967 7001_ $$0P:(DE-HGF)0$$aScapozza, L.$$b3
000030967 7001_ $$0P:(DE-Juel1)128943$$aBott, M.$$b4$$uFZJ
000030967 773__ $$0PERI:(DE-600)1472258-6$$a10.1021/bi0340595$$gVol. 42, p. 5917 - 5924$$p5917 - 5924$$q42<5917 - 5924$$tBiochemistry$$v42$$x0006-2960$$y2003
000030967 8567_ $$uhttp://hdl.handle.net/2128/2417$$uhttp://dx.doi.org/10.1021/bi0340595
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