Probing catalytically essential domain orientation in histidine kinase EnvZ by targeted disulfide crosslinking

Sheng Jian Cai, Ahmad Khorchid, Mitsuhiko Ikura, Masayori Inouye

Research output: Contribution to journalArticlepeer-review

20 Scopus citations

Abstract

EnvZ, a dimeric transmembrane histidine kinase, belongs to the family of His-Asp phosphorelay signal transduction systems. The cytoplasmic kinase domain of EnvZ can be dissected into two independently functioning domains, A and B, whose NMR solution structures have been individually determined. Here, we examined the topological arrangement of these two domains in the EnvZ dimer, a structure that is key to understanding the mechanism underlying the autophosphorylation activity of the kinase. A series of cysteine substitution mutants were constructed to test the feasibility of chemical crosslinking between the two domains. These crosslinking data demonstrate that helix I of domain A of one subunit in the EnvZc dimer is in close proximity to domain B of the other subunit in the same dimer, while helix II of domain A of one subunit interacts with domain B of the same subunit in the EnvZc dimer. This is the first demonstration of the topological arrangement between the central dimerization domain containing the active center His residues (domain A) and the ATP-binding catalysis assisting domain (domain B) in a class I histidine kinase.

Original languageEnglish (US)
Pages (from-to)409-418
Number of pages10
JournalJournal of molecular biology
Volume328
Issue number2
DOIs
StatePublished - Apr 25 2003

All Science Journal Classification (ASJC) codes

  • Structural Biology
  • Molecular Biology

Keywords

  • Disulfide crosslinking
  • Histidine kinase
  • Osmoregulation
  • Signal transduction
  • Structure modeling

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