Structural and functional studies of the HAMP domain of EnvZ, an osmosensing transmembrane histidine kinase in Escherichia coli

Ryuta Kishii, Liliana Falzon, Takeshi Yoshida, Hiroshi Kobayashi, Masayori Inouye

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26 Scopus citations

Abstract

The HAMP domain plays an essential role in signal transduction not only in histidine kinase but also in a number of other signal-transducing receptor proteins. Here we expressed the EnvZ HAMP domain (Arg180-Thr 235) with the R218K mutation (termed LRK) or with L RK connected with domain A (Arg180-Arg289) (termed LARK) of EnvZ, an osmosensing transmembrane histidine kinase in Escherichia coli, by fusing it with protein S. The LRK and LA RK proteins were purified after removing protein S. The CD analysis of the isolated L protein revealed that it consists of a random structure or is unstructured. This suggests that the EnvZ HAMP domain by itself is unable to form a stable structure and that this structural fragility may be important for its role in signal transduction. Interestingly the substitution of Ala 193 in the EnvZ HAMP domain with valine or leucine in Tez1A1, a chimeric protein of Tar and EnvZ, caused a constitutive OmpC phenotype. The CD analysis of LARK(A193L) revealed that this mutated HAMP domain possesses considerable secondary structures and that the thermostability of this entire LARK(A193L) became substantially lower than that of LA RK or just domain A, indicating that the structure of the HAMP domain with the A193L mutation affects the stability of downstream domain A. This results in cooperative thermodenaturation of domain A with the mutated HAMP domain. These results are discussed in light of the recently solved NMR structure of the HAMP domain from a thermophilic bacterium (Hulko, M., Berndt, F., Gruber, M., Linder, J. U., Truffault, V., Schultz, A., Martin, J., Schultz, J. E., Lupas, A. N., and Coles, M. (2006) Cell 126, 929-940).

Original languageEnglish (US)
Pages (from-to)26401-26408
Number of pages8
JournalJournal of Biological Chemistry
Volume282
Issue number36
DOIs
StatePublished - Sep 7 2007

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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