The turgor sensor KdpD of Escherichia coli is a homodimer
- Publication type:
- Journal article
- Metadata:
-
- Autoren
- R Heermann
- K Altendorf
- K Jung
- Autoren-URL
- https://www.webofscience.com/api/gateway?GWVersion=2&SrcApp=fis-test-1&SrcAuth=WosAPI&KeyUT=WOS:000077781500010&DestLinkType=FullRecord&DestApp=WOS_CPL
- DOI
- 10.1016/S0005-2736(98)00181-3
- eISSN
- 1879-2642
- Externe Identifier
- Clarivate Analytics Document Solution ID: 152MN
- PubMed Identifier: 9858704
- ISSN
- 0005-2736
- Ausgabe der Veröffentlichung
- 1
- Zeitschrift
- BIOCHIMICA ET BIOPHYSICA ACTA-BIOMEMBRANES
- Schlüsselwörter
- signal transduction
- sensor kinase
- dimer
- transphosphorylation
- Paginierung
- 114 - 124
- Datum der Veröffentlichung
- 1998
- Status
- Published
- Titel
- The turgor sensor KdpD of <i>Escherichia coli</i> is a homodimer
- Sub types
- Article
- Ausgabe der Zeitschrift
- 1415
Data source: Web of Science (Lite)
- Other metadata sources:
-
- Autoren
- Ralf Heermann
- Karlheinz Altendorf
- Kirsten Jung
- DOI
- 10.1016/s0005-2736(98)00181-3
- ISSN
- 0005-2736
- Ausgabe der Veröffentlichung
- 1
- Zeitschrift
- Biochimica et Biophysica Acta (BBA) - Biomembranes
- Sprache
- en
- Paginierung
- 114 - 124
- Datum der Veröffentlichung
- 1998
- Status
- Published
- Herausgeber
- Elsevier BV
- Herausgeber URL
- http://dx.doi.org/10.1016/s0005-2736(98)00181-3
- Datum der Datenerfassung
- 2021
- Titel
- The turgor sensor KdpD of Escherichia coli is a homodimer
- Ausgabe der Zeitschrift
- 1415
Data source: Crossref
- Abstract
- Escherichia coli responds to K+-limitation or high osmolarity by induction of the kdpFABC operon coding for the high affinity K+-translocating KdpFABC complex. Expression of the corresponding operon is controlled by the membrane-bound sensor kinase KdpD and the cytoplasmic response regulator KdpE. Here, we examine the oligomeric state of KdpD. KdpD-His673-->Gln and KdpD-Asn788-->Asp are kinase inactive. When the corresponding genes are coexpressed, the resulting KdpD protein regains kinase activity in vitro, suggesting that the functional state of KdpD is at least a dimer and that the kinase reaction is a result of a trans-phosphorylation between two monomers. Furthermore, coexpression of kdpD-6His and kdpD-(Delta128-391) leads to stable heterooligomers that can bind to Ni-NTA agarose and that are coeluted. Purified and solubilized KdpD-6His has been electrophoresed in blue native polyacrylamide gels (BN-PAGE), and unphosphorylated and phosphorylated KdpD resulted in the same band pattern suggesting that the oligomeric state of KdpD does not change upon phosphorylation. In addition, determination of the molecular masses of KdpD-6His and KdpD-6His approximately 32P by gel filtration reveals a value of 245 kDa for both forms of the protein. The Stokes radius is determined to be 5.4 nm. Sucrose gradient sedimentation analysis of KdpD-6His results in a molecular mass of 289 kDa. The calculated molecular mass of a KdpD-6His monomer is 99.6 kDa. Considering the detergent bound to KdpD the obtained data reveal that KdpD is a homodimer and there is no change in the oligomeric state upon activation. Crosslinking experiments with single Cys KdpD molecules indicate that there is a close contact between the monomers in the transmitter as well as in transmembrane domain 1. BN-PAGE of solubilized and purified KdpD-6His devoid of Cys residues demonstrates that Cys residues do not contribute to the stabilization of the dimer.
- Addresses
- Universität Osnabrück, Fachbereich Biologie/Chemie, Abteilung Mikrobiologie, D-49069 Osnabrück, Germany.
- Autoren
- R Heermann
- K Altendorf
- K Jung
- DOI
- 10.1016/s0005-2736(98)00181-3
- eISSN
- 1878-2434
- Externe Identifier
- PubMed Identifier: 9858704
- Open access
- false
- ISSN
- 0006-3002
- Ausgabe der Veröffentlichung
- 1
- Zeitschrift
- Biochimica et biophysica acta
- Schlüsselwörter
- Escherichia coli
- Phosphotransferases
- Protein Kinases
- Bacterial Proteins
- Escherichia coli Proteins
- Chromatography, Gel
- Electrophoresis, Polyacrylamide Gel
- Mutagenesis, Site-Directed
- Biosensing Techniques
- Dimerization
- Phosphorylation
- Molecular Weight
- Sprache
- eng
- Medium
- Paginierung
- 114 - 124
- Datum der Veröffentlichung
- 1998
- Status
- Published
- Datum der Datenerfassung
- 1998
- Titel
- The turgor sensor KdpD of Escherichia coli is a homodimer.
- Sub types
- Research Support, Non-U.S. Gov't
- Journal Article
- Ausgabe der Zeitschrift
- 1415
Data source: Europe PubMed Central
- Abstract
- Escherichia coli responds to K+-limitation or high osmolarity by induction of the kdpFABC operon coding for the high affinity K+-translocating KdpFABC complex. Expression of the corresponding operon is controlled by the membrane-bound sensor kinase KdpD and the cytoplasmic response regulator KdpE. Here, we examine the oligomeric state of KdpD. KdpD-His673-->Gln and KdpD-Asn788-->Asp are kinase inactive. When the corresponding genes are coexpressed, the resulting KdpD protein regains kinase activity in vitro, suggesting that the functional state of KdpD is at least a dimer and that the kinase reaction is a result of a trans-phosphorylation between two monomers. Furthermore, coexpression of kdpD-6His and kdpD-(Delta128-391) leads to stable heterooligomers that can bind to Ni-NTA agarose and that are coeluted. Purified and solubilized KdpD-6His has been electrophoresed in blue native polyacrylamide gels (BN-PAGE), and unphosphorylated and phosphorylated KdpD resulted in the same band pattern suggesting that the oligomeric state of KdpD does not change upon phosphorylation. In addition, determination of the molecular masses of KdpD-6His and KdpD-6His approximately 32P by gel filtration reveals a value of 245 kDa for both forms of the protein. The Stokes radius is determined to be 5.4 nm. Sucrose gradient sedimentation analysis of KdpD-6His results in a molecular mass of 289 kDa. The calculated molecular mass of a KdpD-6His monomer is 99.6 kDa. Considering the detergent bound to KdpD the obtained data reveal that KdpD is a homodimer and there is no change in the oligomeric state upon activation. Crosslinking experiments with single Cys KdpD molecules indicate that there is a close contact between the monomers in the transmitter as well as in transmembrane domain 1. BN-PAGE of solubilized and purified KdpD-6His devoid of Cys residues demonstrates that Cys residues do not contribute to the stabilization of the dimer.
- Autoren
- R Heermann
- K Altendorf
- K Jung
- Autoren-URL
- https://www.ncbi.nlm.nih.gov/pubmed/9858704
- DOI
- 10.1016/s0005-2736(98)00181-3
- ISSN
- 0006-3002
- Ausgabe der Veröffentlichung
- 1
- Zeitschrift
- Biochim Biophys Acta
- Schlüsselwörter
- Bacterial Proteins
- Biosensing Techniques
- Chromatography, Gel
- Dimerization
- Electrophoresis, Polyacrylamide Gel
- Escherichia coli
- Escherichia coli Proteins
- Molecular Weight
- Mutagenesis, Site-Directed
- Phosphorylation
- Phosphotransferases
- Protein Kinases
- Sprache
- eng
- Country
- Netherlands
- Paginierung
- 114 - 124
- PII
- S0005-2736(98)00181-3
- Datum der Veröffentlichung
- 1998
- Status
- Published
- Datum, an dem der Datensatz öffentlich gemacht wurde
- 1999
- Titel
- The turgor sensor KdpD of Escherichia coli is a homodimer.
- Sub types
- Journal Article
- Research Support, Non-U.S. Gov't
- Ausgabe der Zeitschrift
- 1415
Data source: PubMed
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