title: Crystal structure of the VapBC-15 complex from Mycobacterium tuberculosis reveals a two-metal ion dependent PIN-domain ribonuclease and a variable mode of toxin-antitoxin assembly.
abstract: Although PIN (PilT N-terminal)-domain proteins are known to have ribonuclease activity, their specific mechanism of action remains unknown. VapCs form a family of ribonucleases that possess a PIN-domain assembly and are known as toxins. The activities of VapCs are impaired by VapB antitoxins. Here we present the crystal structure of the VapBC-15 toxin-antitoxin complex from Mycobacterium tuberculosis determined to 2.1Å resolution. The VapB-15 and VapC-15 components assemble into one heterotetramer (VapB2C2) and two heterotrimers (VapBC2) in each asymmetric unit of the crystal. The active site of VapC-15 toxin consists of a cluster of acidic amino acid residues and two divalent metal ions, forming a well organised ribonuclease active site. The distribution of the catalytic-site residues of the VapC-15 toxin is similar to that of T4 RNase H and of Methanococcus jannaschii FEN-1, providing strong evidence that these three proteins share a similar mechanism of activity. The presence of both VapB2C2 and VapBC2 emphasizes the fact that the same antitoxin can bind the toxin in 1:1 and 1:2 ratios. The crystal structure determination of the VapBC-15 complex reveals for the first time a PIN-domain ribonuclease protein that shows two metal ions at the active site and a variable mode of toxin-antitoxin assembly. The structure further shows that VapB-15 antitoxin binds to the same groove meant for the binding of putative substrate (RNA), resulting in the inhibition of VapC-15's toxicity.
authors.affiliation: Department of Biophysics, All India Institute of Medical Sciences, New Delhi, India.
authors.forename: Uddipan
authors.initials: U
authors.lastname: Das
authors.affiliation: European Molecular Biology Laboratory, Hamburg, Germany.
authors.forename: Vivian
authors.initials: V
authors.lastname: Pogenberg
authors.affiliation: Centre for Structural Systems Biology (CSSB), DESY Campus, Hamburg, Germany and Forschungszentrum Jülich GmbH, Juelich, Germany.
authors.forename: Udaya Kumar Tiruttani
authors.initials: UK
authors.lastname: Subhramanyam
authors.affiliation: European Molecular Biology Laboratory, Hamburg, Germany.
authors.forename: Matthias
authors.initials: M
authors.lastname: Wilmanns
authors.affiliation: School of Life Sciences, Jawaharlal Nehru University, New Delhi, India. Electronic address: sgourinath@mail.jnu.ac.in.
authors.forename: Samudrala
authors.initials: S
authors.lastname: Gourinath
authors.affiliation: Department of Biophysics, All India Institute of Medical Sciences, New Delhi, India. Electronic address: srini@aiims.ac.in.
authors.forename: Alagiri
authors.initials: A
authors.lastname: Srinivasan
mesh_terms: D000992:Antitoxins / Q000378:metabolism*; D001426:Bacterial Proteins / Q000378:metabolism*; D001427:Bacterial Toxins / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D008562:Membrane Glycoproteins / Q000378:metabolism*; D008958:Models, Molecular; D009169:Mycobacterium tuberculosis / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D012260:Ribonucleases / Q000378:metabolism*; D014961:X-Ray Diffraction
chemical_list: D000992:Antitoxins; D001426:Bacterial Proteins; D001427:Bacterial Toxins; D004268:DNA-Binding Proteins; D008562:Membrane Glycoproteins; C077517:VapB protein, Bacteria; D012260:Ribonucleases
keywords: Mycobacterium tuberculosis; Toxin; Toxin–antitoxin; VapB-15; VapBC-15; VapC-15; X-ray crystallography