Literature summary for 2.7.13.3 extracted from

Affandi, T.; Issaian, A.V.; McEvoy, M.M.
The structure of the periplasmic sensor domain of the histidine kinase CusS shows unusual metal ion coordination at the dimeric interface (2016), Biochemistry, 55, 5296-5306.

Cloned(Commentary)

Cloned (Comment)	Organism
recombinant expression of His6-tagged wild-type and mutant enzymes in Escherichia coli. The expression of CusS from a plasmid does not result in the same growth phenotype on copper-containing media as the strain that expresses chromosomal CusS	Escherichia coli

Crystallization (Commentary)

Crystallization (Comment)	Organism
purified recombinant Ag(I)-bound periplasmic sensor domain, Ag(I)-CusS(39-187), sitting drop vapor diffusion method, mixing of 200 nl of 7-8 mg/ml protein in 25 mM MES, pH 6.0, with 200 nl of precipitant solution containing 100 mM Tris-HCl, pH 8.5, 200 mM ammonium acetate, and 25% PEG 3350, 4°C, equilibration against 1 ml precipitant solution, method optimization, X-ray diffraction structure determination and analysis, single anomalous diffraction (SAD) method at 2.15 A resolution. There are four Ag(I)-CusS(39-187) molecules in the asymmetric unit that interact to form two homodimers	Escherichia coli

Crystallization (Comment)

Organism

purified recombinant Ag(I)-bound periplasmic sensor domain, Ag(I)-CusS(39-187), sitting drop vapor diffusion method, mixing of 200 nl of 7-8 mg/ml protein in 25 mM MES, pH 6.0, with 200 nl of precipitant solution containing 100 mM Tris-HCl, pH 8.5, 200 mM ammonium acetate, and 25% PEG 3350, 4°C, equilibration against 1 ml precipitant solution, method optimization, X-ray diffraction structure determination and analysis, single anomalous diffraction (SAD) method at 2.15 A resolution. There are four Ag(I)-CusS(39-187) molecules in the asymmetric unit that interact to form two homodimers

Escherichia coli

Protein Variants

Protein Variants	Comment	Organism
F43I	site-directed mutagenesis, the mutation results in a slightly reduced growth defect compared to the construct in which all interface binding site residues are mutated	Escherichia coli
H176A	site-directed mutagenesis, the mutation results in the same growth defects as the construct in which all interface binding site residues are mutated	Escherichia coli
H42A	site-directed mutagenesis, the mutation results in the same growth defects as the construct in which all interface binding site residues are mutated	Escherichia coli
additional information	construction of a DELTAcusS enzyme nockout mutant in an Escherichia coli strain in which the chromosomal copy of the gene for the multicopper oxidase CueO has been deleted (DELTAcueO). CueO converts Cu(I) to Cu(II) and therefore the use of the strain with the cueO deletion allows the observation of a growth phenotype even under aerobic conditions. The chromosomal copy of cusS is deleted from this background strain to allow characterization of CusS variants expressed from a plasmid. Partial functional complementation ofthe mutant strain with lasmid-expressed wild-type enzyme. The mutations do not significantly affect the relative expression levels of CusS	Escherichia coli

Inhibitors

Inhibitors	Comment	Organism	Structure
Cu2+	the wild-type strain experiences copper susceptibility at 2.75 mM CuSO4, whereas DELTAcusS mutant cells show a phenotype at lower concentrations	Escherichia coli

Localization

Localization	Comment	Organism	GeneOntology No.	Textmining
membrane	CusS is a membrane-associated histidine kinase with a periplasmic sensor domain connected to the cytoplasmic ATP-binding and catalytic domains through two transmembrane helices	Escherichia coli	16020	-
additional information	the structure of the periplasmic sensor domain of the histidine kinase CusS shows unusual metal ion coordination at the dimeric interface	Escherichia coli	-	-

Metals/Ions

Metals/Ions	Comment	Organism
Ag+	CusS forms a homodimer with four Ag(I) binding sites per dimeric complex, binding site structure, overview	Escherichia coli
Mg2+	required	Escherichia coli
additional information	the structure of the periplasmic sensor domain of the histidine kinase CusS shows unusual metal ion coordination at the dimeric interface. Two symmetric metal binding sites are found at the dimeric interface, which are each formed by two histidines and one phenylalanine with an unusual cation-Pi interaction, the other metal ion binding sites are in a non-conserved region within each monomer. The metal ion binding site at the dimer interface is more important for function. Metal-induced dimerization results in increases in kinase activity in the cytoplasmic domains of CusS. Functional roles of the metal binding sites in CusS, overview	Escherichia coli

Organism

Organism	UniProt	Comment	Textmining
Escherichia coli	P77485	-	-

Posttranslational Modification

Posttranslational Modification	Comment	Organism
phosphoprotein	ATP-dependent autophosphorylation occurs at a conserved His residue in the kinase core. The phosphoryl group is then transferred from the histidine kinase to a conserved Asp residue on the N-terminal receiver domain of the response regulator, which in turn activates the C-terminal effector domain to mediate an adaptive response. The phosphorylated response regulator activates transcription of genes that respond to the environmental stimulus	Escherichia coli

Subunits

Subunits	Comment	Organism
dimer	the overall structure of Ag(I)-CusS(39-187) is a mixed alpha/beta fold with a PAS-like topology. The central five-stranded anti-parallel beta-sheet is flanked by alpha-helices on either side. The structure begins with a long N-terminal alpha-helix and ends with a short C-terminal alpha-helix that both lie on the same side of the sheet, The structure of the periplasmic sensor domain of the histidine kinase CusS shows unusual metal ion coordination at the dimeric interface, enzyme CusS forms a homodimer with four Ag(I) binding sites per dimeric complex. Metal-induced dimerization results in increases in kinase activity in the cytoplasmic domains of CusS. There are four Ag(I)-CusS(39-187) molecules in the asymmetric unit that interact to form two homodimers	Escherichia coli

Synonyms

Synonyms	Comment	Organism
CusS	-	Escherichia coli

Cofactor

Cofactor	Comment	Organism	Structure
ATP	-	Escherichia coli

General Information

General Information	Comment	Organism
malfunction	the wild-type strain experiences copper susceptibility at 2.75 mM CuSO4, whereas DELTAcusS mutant cells show a phenotype at lower concentrations. The growth defects observed in the DELTAcusS strain can be partially rescued by expression of intact CusS from a plasmid	Escherichia coli
physiological function	in bacteria, two-component systems act as signaling systems to respond to environmental stimuli. Two-component systems generally consist of a sensor histidine kinase and a response regulator, which work together through histidyl-aspartyl phospho-relay to result in gene regulation. One of the two-component systems in Escherichia coli, CusS-CusR, is known to induce expression of cusCFBA genes under increased periplasmic Cu(I) and Ag(I) concentrations to help maintain metal ion homeostasis. CusS senses increasing metal ion concentrations and activates CusR. ATP-dependent auto-phosphorylation occurs at a conserved His residue in the kinase core. The phosphoryl group is then transferred from the histidine kinase to a conserved Asp residue on the N-terminal receiver domain of the response regulator, which in turn activates the C-terminal effector domain to mediate an adaptive response. The phosphorylated response regulator activates transcription of genes that respond to the environmental stimulus. Analysis of the molecular role of CusS in Cu(I)/Ag(I) resistance and the mechanism of CusS signal transduction in Escherichia coli, overview. In response to periplasmic Ag(I) and Cu(I), CusS upregulates the genes to express the CusCBA Cu(I)/Ag(I) efflux pump to remove excess metal ions from the cell	Escherichia coli