Biochemical characterization of the tandem HAMP domain from Natronomonas pharaonis as an intraprotein signal transducer
Autor: | Janani Natarajan, Joachim E. Schultz, Anita Schultz, Ursula Kurz |
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Rok vydání: | 2014 |
Předmět: |
Archaeal Proteins
Mutant Mutant Chimeric Proteins Biology Biochemistry Cyclase HAMP domain Adenylyl cyclase Serine chemistry.chemical_compound Bacterial Proteins Escherichia coli Amino Acid Sequence Molecular Biology Halobacteriaceae Membrane Proteins Cell Biology Cell biology Protein Structure Tertiary chemistry HAMP Signal transduction Cyclase activity Adenylyl Cyclases Signal Transduction |
Zdroj: | The FEBS journal. 281(14) |
ISSN: | 1742-4658 |
Popis: | Available structures of HAMP domains suggest rotation as one potential mechanism in intraprotein signal transduction. It has been proposed that in poly-HAMP modules the signal sign is inverted with each additional HAMP. We examined signal transduction through the HAMP tandem domain from the phototaxis transducer of the halophilic archaeon Natronomonas pharaonis in membrane-bound chimeras consisting of the Escherichia coli chemotaxis receptor for serine, Tsr, as an input and the mycobacterial adenylyl cyclase Rv3645 as an output domain, i.e. the basic chimera was 'Tsr-NpHAMP tandem-Rv3645 cyclase'. Neither of the NpHAMP units alone nor the NpHAMP tandem transduced a serine signal. After five targeted point mutations in the first α-helix of NpHAMP1 , the non-functional NpHAMP modules combined into a functional HAMP tandem. 1 mm serine significantly inhibited cyclase activity (-35%; IC50 = 30 μm) in disagreement with the structure-based predictions. Surprisingly, replacement of NpAS11 in the tandem by the respective AS1 from HAMPT sr resulted in signal inversion, i.e. serine activated cyclase (+129%; EC50 = 10 μm). Examination of 48 mutants of AS11 in the HAMP tandem including two residues of a putative N-terminal control cable identified five residues in NpAS11 which probably define different ground states of the output domain and thus affect the sign of signal output. The data question the predicted HAMP rotation as the predominant mechanism of intraprotein signal transduction and point to as yet unrecognized conformational motions of HAMP domains in intraprotein signaling. |
Databáze: | OpenAIRE |
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