Functional and Structural Characterization of EnvZ, an Osmosensing Histidine Kinase of E. coli

Research output: Chapter in Book/Report/Conference proceedingChapter

Abstract

EnvZ is an osmosensing histidine kinase located in the inner membrane, and one of the most extensively studied Escherichia coli histidine kinases. Because of its structural complexity, functional and structural studies have been quite challenging. It is a multidomain transmembrane protein consisting of 450 amino acid residues. In addition, it must form a dimer to function as a histidine kinase like all the other histidine kinases. EnvZ consists of the 115-residue periplasmic domain, two transmembrane domains (TM1 and TM2), and the cytoplasmic domain consisting of the 43-residue linker (HAMP) domain and the 228-residue kinase domain. It has been shown that the kinase domain of EnvZ, responsible for its enzymatic activities, contains all of the conserved regions of histidine kinases such as H, F, N, G1, G2, and G3 boxes. Therefore, the 271-residue cytoplasmic domain of EnvZ (termed EnvZc) has been used as a model system to establish fundamental characteristics of histidine kinases. The DNA fragment encoding EnvZc was cloned in pET vector and EnvZc was expressed and purified. It is highly soluble and retains all the enzymatic activities of EnvZ. We demonstrated that it consists of two functional domains, domain A and domain B. NMR spectroscopic studies of these two domains revealed, for the first time, the structure of a histidine kinase. Domain A is responsible for dimerization of EnvZc forming a four-helical bundle containing two α-helical hairpin structures, while domain B is a monomer and has an ATP-binding pocket formed by regions conserved among the histidine kinases. In this chapter, we describe functional and structural studies of EnvZc, which can be applied to characterize other histidine kinases.

Original languageEnglish (US)
Title of host publicationTwo Component Signaling Systems, Part B
PublisherAcademic Press Inc.
Pages184-202
Number of pages19
Volume423
DOIs
StatePublished - Jan 1 2007

Publication series

NameMethods in Enzymology
Volume423

Fingerprint

Histidine
Escherichia coli
Phosphotransferases
Histidine Kinase
Dimerization
Dimers
Adenosine Triphosphate
Amino Acids
Monomers
Nuclear magnetic resonance
Membranes
DNA

All Science Journal Classification (ASJC) codes

  • Molecular Biology
  • Biochemistry

Cite this

Yoshida, T., Phadtare, S., & Inouye, M. (2007). Functional and Structural Characterization of EnvZ, an Osmosensing Histidine Kinase of E. coli. In Two Component Signaling Systems, Part B (Vol. 423, pp. 184-202). (Methods in Enzymology; Vol. 423). Academic Press Inc.. https://doi.org/10.1016/S0076-6879(07)23008-3
Yoshida, Takeshi ; Phadtare, Sangita ; Inouye, Masayori. / Functional and Structural Characterization of EnvZ, an Osmosensing Histidine Kinase of E. coli. Two Component Signaling Systems, Part B. Vol. 423 Academic Press Inc., 2007. pp. 184-202 (Methods in Enzymology).
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Yoshida, T, Phadtare, S & Inouye, M 2007, Functional and Structural Characterization of EnvZ, an Osmosensing Histidine Kinase of E. coli. in Two Component Signaling Systems, Part B. vol. 423, Methods in Enzymology, vol. 423, Academic Press Inc., pp. 184-202. https://doi.org/10.1016/S0076-6879(07)23008-3

Functional and Structural Characterization of EnvZ, an Osmosensing Histidine Kinase of E. coli. / Yoshida, Takeshi; Phadtare, Sangita; Inouye, Masayori.

Two Component Signaling Systems, Part B. Vol. 423 Academic Press Inc., 2007. p. 184-202 (Methods in Enzymology; Vol. 423).

Research output: Chapter in Book/Report/Conference proceedingChapter

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Yoshida T, Phadtare S, Inouye M. Functional and Structural Characterization of EnvZ, an Osmosensing Histidine Kinase of E. coli. In Two Component Signaling Systems, Part B. Vol. 423. Academic Press Inc. 2007. p. 184-202. (Methods in Enzymology). https://doi.org/10.1016/S0076-6879(07)23008-3