Regulatory Implications of Structural Changes in Tyr201 of the Oxygen Sensor Protein FixL

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• 作者：Takeo Yamawaki ; Haruto Ishikawa ; Misao Mizuno ; Hiro Nakamura ; Yoshitsugu Shiro ; Yasuhisa Mizutani
• 刊名：Biochemistry
• 出版年：2016
• 出版时间：July 26, 2016
• 年：2016
• 卷：55
• 期：29
• 页码：4027-4035
• 全文大小：477K
• 年卷期：0
• ISSN：1520-4995

文摘

FixL is a heme-based oxygen-sensing histidine kinase that induces the expression of nitrogen fixation genes under hypoxic conditions. Oxygen dissociation from heme iron in the sensor domain of FixL initiates protein conformational changes that are transmitted to the histidine kinase domain, activating autophosphorylation activity. Conversely, oxygen binding inhibits FixL kinase activity. It is essential to elucidate the changes that occur in the protein structure upon this oxygen dissociation for understanding of the allosteric transduction mechanism. We measured ultraviolet resonance Raman spectra of FixL and its mutants for deoxy, oxy, and carbonmonoxy forms to examine the changes in protein structure upon oxygen dissociation. The observed spectral changes indicated that Tyr201 and its neighboring residues undergo structural changes upon oxygen dissociation. Kinase assays showed that substitution of Tyr201 significantly decreased the inhibition of kinase activity upon oxygen binding. These data mean that weakening of the hydrogen bond of Tyr201 that is induced by oxygen dissociation is essential for inhibition of kinase activity. We also observed spectral changes in Tyr residues in the kinase domain upon oxygen dissociation from FixL, which is the first observation of oxygen-dependent structural changes in the kinase domain of FixL. The observed structural changes support the allosteric transduction pathway of FixL which we proposed previously [Yano, S., Ishikawa, H., Mizuno, M., Nakamura, H., Shiro, Y., and Mizutani, Y.x"><x xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:ACS="http://namespace.acs.org/2008/acs" xmlns:xsi="http://www.w3.org/2001/XMLSchema-instance" xml:space="preserve"> (x>2013x"><x xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:ACS="http://namespace.acs.org/2008/acs" xmlns:xsi="http://www.w3.org/2001/XMLSchema-instance" xml:space="preserve">) x> J. Phys. Chem. B117x"><x xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:ACS="http://namespace.acs.org/2008/acs" xmlns:xsi="http://www.w3.org/2001/XMLSchema-instance" xml:space="preserve">, x>15786−15791].