The KdpF subunit is part of the K(+)-translocating Kdp complex of Escherichia coli and is responsible for stabilization of the complex in vitro

J Biol Chem. 1999 Dec 31;274(53):37901-7. doi: 10.1074/jbc.274.53.37901.

Abstract

The kdpABC operon codes for the high affinity K(+)-translocating Kdp complex (P-type ATPase) of Escherichia coli. Upon expression of this operon in minicells, a so far unrecognized small hydrophobic polypeptide, KdpF, could be identified on high resolution SDS-polyacrylamide gels in addition to the subunits KdpA, KdpB, and KdpC. Furthermore, it could be demonstrated that KdpF remains associated with the purified complex. As determined by mass spectrometry, this peptide is present in its formylated form and has a molecular mass of 3100 Da. KdpF is not essential for growth on low K(+) (0.1 mM) medium, as shown by deletion analysis of kdpF, but proved to be indispensable for a functional enzyme complex in vitro. In the absence of KdpF, the ATPase activity of the membrane-bound Kdp complex was almost indistinguishable from that of the wild type. In contrast, the purified detergent-solubilized enzyme complex showed a dramatic decrease in enzymatic activity. However, addition of purified KdpF to the KdpABC complex restored the activity up to wild type level. It is interesting to note that the addition of high amounts of E. coli lipids had a similar effect. Although KdpF is not essential for the function of the Kdp complex in vivo, it is part of the complex and functions as a stabilizing element in vitro. The corresponding operon should now be referred to as kdpFABC.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenosine Triphosphatases / chemistry
  • Adenosine Triphosphatases / genetics
  • Adenosine Triphosphatases / isolation & purification
  • Adenosine Triphosphatases / metabolism*
  • Amino Acid Sequence
  • Base Sequence
  • Carrier Proteins / chemistry
  • Carrier Proteins / genetics
  • Carrier Proteins / isolation & purification
  • Carrier Proteins / metabolism*
  • Cation Transport Proteins*
  • Chromatography, Ion Exchange
  • Codon, Initiator
  • Enzyme Activation
  • Enzyme Stability
  • Escherichia coli / growth & development
  • Escherichia coli / metabolism*
  • Escherichia coli Proteins*
  • Ion Transport
  • Molecular Sequence Data
  • Mutagenesis, Site-Directed
  • Operon
  • Potassium / metabolism*

Substances

  • Carrier Proteins
  • Cation Transport Proteins
  • Codon, Initiator
  • Escherichia coli Proteins
  • Adenosine Triphosphatases
  • potassium translocating Kdp-ATPase, E coli
  • Potassium