tailieunhanh - Báo cáo khoa hoc : The role of the GAF and central domains of the transcriptional activator VnfA in Azotobacter vinelandii

VnfA is a transcriptional activator that is required for the expression of the structural genes encoding nitrogenase-2 in Azotobacter vinelandii. VnfA consists of three domains: an N-terminal regulatory domain termed GAF, including a Cys-rich motif; a central domain from the AAA+ family; and a C-terminal domain for DNA binding. | IFEBS Journal The role of the GAF and central domains of the transcriptional activator VnfA in Azotobacter vinelandii Kyohei Yoshimitsu1 Nobuyuki Takatani2 Yukio Miura1 Yoshihito Watanabe3 and Hiroshi Nakajima1 1 Department of Chemistry Graduate Schoolof Science Nagoya University Japan 2 Graduate Schoolof BioagriculturalScience Nagoya University Japan 3 Research Center of Materials Science Nagoya University Japan Keywords Azotobacter vinelandii iron-sulfur cluster nitrogenase NTPase activity transcriptional regulator Correspondence H. Nakajima Department of Chemistry Graduate School of Science Nagoya University Furo-cho Chikusa-ku Nagoya 464-8602 Japan Fax 81 52 789 3557 Tel 81 52 789 2953 E-mail hnakajima@. Received 19 April 2011 revised 6 July 2011 accepted 8 July 2011 doi VnfA is a transcriptional activator that is required for the expression of the structural genes encoding nitrogenase-2 in Azotobacter vinelandii. VnfA consists of three domains an N-terminal regulatory domain termed GAF including a Cys-rich motif a central domain from the AAA family and a C-terminal domain for DNA binding. Previously we reported that transcriptionally active VnfA harboring an Fe-S cluster presumably of the 3Fe-4S type as a prosthetic group and the Cys-rich motif were possibly associated with coordination of the Fe-S cluster. In the present study we have investigated the roles of the GAF and central domains in the regulatory function of VnfA using truncated variants AN15VnfA and AGAFVnfA that lack the N-terminal 15 residues and whole GAF domain respectively and GAFVnfA consisting of only the GAF domain. AN15VnfA and AGAFVnfA lost the ability to bind the Fe-S cluster whereas GAFVnfA was still able to bind to the cluster consistent with the hypothesis that the Cys-rich motif is essential for Fe-S cluster binding. The GAF domain showed an inhibitory effect on the transcriptional activity of VnfA which was reversed in the .