Amino acid replacements in transmembrane domain 1 influence osmosensing but not K+ sensing by the sensor kinase KdpD of Escherichia coli.
Expression of the kdpFABC operon coding for the high affinity K+ -translocating KdpFABC complex of Escherichia coli is induced by K+ limitation or high osmolality. This process is controlled by the sensor kinase/response regulator system KdpD/KdpE. To study the importance of the transmembrane domains of KdpD for stimulus perception, each amino acid residue of the transmembrane domain 1 and Asp-424 of the adjacent periplasmic loop were replaced with Cys in a KdpD derivative devoid of native Cys residues. In vivo analysis of KdpD proteins with a single Cys residue revealed that 14 out of 18 amino acid replacements caused an altered response towards an osmotic upshift imposed by NaCl, whereby only four replacements also altered the response towards changes in the K+ concentration. The in vitro activities of most of the KdpD derivatives were in the range of KdpD devoid of native Cys residues. The results reveal that the osmosensing and K+ -sensing properties of KdpD can be dissected. Furthermore, the data support the hypothesis that osmosensing involves amino acid residues of the transmembrane domains.
Stallkamp
Iris
I
Universität Osnabrück, Fachbereich Biologie/Chemie, Abteilung Mikrobiologie, 49069 Osnabrück, Germany.
Altendorf
Karlheinz
K
Jung
Kirsten
K
D019943:Amino Acid Substitution; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D003545:Cysteine / Q000235:genetics / Q000378:metabolism; D004220:Disulfides / Q000032:analysis / Q000737:chemistry; D004926:Escherichia coli / Q000737:chemistry / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D007700:Kinetics; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D008956:Models, Chemical; D016297:Mutagenesis, Site-Directed; D009876:Operon; D011188:Potassium / Q000378:metabolism*; D011494:Protein Kinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D013329:Structure-Activity Relationship
D001426:Bacterial Proteins; D004220:Disulfides; D029968:Escherichia coli Proteins; D008565:Membrane Proteins; C489580:KdpD protein, E coli; D011494:Protein Kinases; C075616:kdpD protein, Bacteria; D003545:Cysteine; D011188:Potassium