Molecular Playground/CsoR and RcnR

spinqualitylabelsall models Cu(I)-bound CsoR (PDB ID: 2HH7) Show:Asymmetric Unit Biological Assembly Drag the structure with the mouse to rotate   Export Animated Image   Print3D One of the CBI Molecules being studied in the University of Massachusetts Amherst Chemistry-Biology Interface Program at UMass Amherst and on display at the Molecular Playground. Introduction Heavy metals such as iron, nickel, copper, and zinc are important cofactors for the functions of many different metalloenzymes. High levels of these heavy metals can also cause damage to cellular components, therefore intracellular levels of metals are tightly regulated within the cell. One of the ways that bacteria can regulate intracellular metal levels is by increasing the amount of metal efflux proteins. CsoR, RcnR and InrS are members of a large family of metal-responsive DNA-binding proteins, all of which regulate the transcription of metal-specific efflux proteins. CsoR is only responsive to the binding of Cu(I); RcnR is responsive to the binding of Ni(II) or Co(II); whereas InrS is only responsive to the binding of Ni(II). CsoR/RcnR Family In Escherichia coli, apo-RcnR blocks the transcription of nickel and cobalt efflux proteins RcnA and RcnB by binding to its promoter region. Although no crystal structure of RcnR is available on the PDB, the Cu(I)-bound CsoR crystal structure from Mycobacterium tuberculosis is available and RcnR is predicted to share a similar fold to CsoR. Upon Ni(II)- or Co(II)-binding, RcnR is released from DNA allowing the transcription of RcnA and RcnB, facilitating the efflux of Ni(II) and Co(II). CsoR has been characterized in Bacillus subtilis and M. tuberculosis to release from the promoter regions of copper-efflux operons upon binding of Cu(I). InrS is another member of this metal-responsive transcriptional family recently found in Synechocystis. InrS regulates cytosolic nickel level by releasing from the promoter regions of nickel-efflux operons upon Ni(II) binding. The analogous functions of CsoR, RcnR and InrS in addition to local sequence similarity (RcnR has 24% identity, 67% similarity, and InrS has 36% idendity and 72% similarity with M. tuberculosis CsoR.) suggests a conserved mode of function in this family of metal-responsive DNA-binding proteins. Structure of CsoR/RcnR Family Tetrameric CsoR binds one Cu(I) per monomer. The protein forms a dimer of dimers with a pore in the tetrameric interface. Each Cu(I) coordinated by two different monomers of a dimer unit. Where one monomer is bound to Cu(I) by His61 and Cys65, the other monomer is bound to the metal by Cys35. Hydrogen bonds in the second coordination sphere residues, Tyr35 and Glu81, with non-coordinating face of His61 stabilize the Cu(I) complex of CsoR. RcnR is also tetrameric and has the same protein-to-metal stoichiometry. Through a combination of site-directed mutagenesis, lacZ activity assays and X-ray absorption spectroscopy (XAS) structural studies it is now known that RcnR uses a different ligand set to bind Ni(II) than it does Co(II). The Ni(II) site is bound by the N-terminal amine, Cys35, and His64. Because the Ni(II) site has an octahedral geometry, there are three metal binding residues whose identity is unknown. However, using XAS and lacZ activity assays it is known that the remaining ligands bind to the Ni(II) using N/O atoms and that there are a total of two histidine ligand bound to the Ni(II), leaving the identity of one unaccounted for. Studies have shown that this histidine is not His3, His60, or His67.The Co(II) site is bound by the the N-terminal amine, His3, Cys35, His64 and possibly His60. The Co(II) site of RcnR also has an octahedral geometry, leaving 1-2 residues whose identity is unknown. XAS reveals that these ligands are bound to Co(II) using N/O atoms and that there are a total of three histidine ligands bound to the Co(II). Studies have shown that His67 is not a Co(II) ligand. In addition, RcnR does not conserved the second coordination sphere hydrogen bond network, implying that there is a distance allosteric mechanism relative to CsoR. InrS conserves all the Cu(I) binding residues identified in M. tuberculosis CsoR, Cys53, Cys82 and His78, which have been shown as Ni(II) ligands. His21 is also important for metal binding, while it remains unclear whether or not His21 is a ligand involved in this square planar metal site. Added to this, InrS also lacks the second coordination sphere hydrogen bond network. Fig. 1: Schematic of known Ni(II) and Co(II) binding residues in the metal sites of RcnR, Cu(I) binding residues in CsoR and Ni(II) binding site in InrS. Figure made with ChemDraw. Research Interests The mechanism of DNA binding of the CsoR/RcnR family of metal-responsive transcriptional regulators is still unknown. Additionally, RcnR has an added level of complexity because it is reponsive to both Ni(II) and Co(II) binding. The Maroney Lab at the University of Massachusetts Amherst is interested in the conformational changes of RcnR induced by DNA-, Ni(II)-, and Co(II)-binding. Identification of the remaining metal binding residues in RcnR is ongoing, in addition to identification of the DNA-binding residues in RcnR, as there is no crystal structure of any member of this family of proteins binding to DNA.

3D structures of copper homeostasis protein

CBI Molecules

Also See

2hh7

Proteopedia Page Contributors and Editors (what is this?)

Heidi Hu, Carolyn Carr, Hsin-Ting Huang, Michal Harel, Alexander Berchansky