An XAS investigation of the nickel site structure in the transcriptional regulator InrS

Abstract

InrS (Internal nickel-responsive Sensor) is a transcriptional repressor of the nickel exporter NrsD and de-represses expression of the exporter upon binding Ni(II) ions. Although a crystal structure of apo-InrS has been reported, no structure of the protein with metal ions bound is available. Herein we report the results of metal site structural investigations of Ni(II) and Cu(II) complexes of InrS using x-ray absorption spectroscopy (XAS) that are complementary to data available from the apo-InrS crystal structure, and are consistent with a planar four-coordinate [Ni(His)2(Cys)2] structure, where the ligands are derived from the side chains of His21, Cys53, His78, and Cys82. Coordination of Cu(II) to InrS forms a nearly identical planar four-coordinate complex that is consistent with a simple replacement of the Ni(II) center by Cu(II).

Molecular Modelling of the Ni(II)-Responsive Synechocystis PCC 6803 Transcriptional Regulator InrS in the Metal Bound Form

“…The alignment between the SyInrS sequence and the other CsoR/RcnR family members revealed that the canonical W-X-Y-Z fingerprint, used to characterize the first coordination shell residues [8,9], corresponds to His21, Cys53, His78, and Cys82 in SyInrS (Figure 1). A subsequent mutagenesis study confirmed the roles of the latter three residues in Ni(II) binding [16], while His21 has been recently confirmed as the fourth Ni(II) ligand both through mutagenesis [17] and X-ray absorption spectroscopy (XAS) data [18]. The latter study, which investigated both Ni(II) and Cu(II) bound SyInrS, also confirmed the presence of a planar four-coordinate [Ni(His) 2 (Cys) 2 ] site and provided Ni(II)-ligands interatomic distances [18].…”

“…A subsequent mutagenesis study confirmed the roles of the latter three residues in Ni(II) binding [16], while His21 has been recently confirmed as the fourth Ni(II) ligand both through mutagenesis [17] and X-ray absorption spectroscopy (XAS) data [18]. The latter study, which investigated both Ni(II) and Cu(II) bound SyInrS, also confirmed the presence of a planar four-coordinate [Ni(His) 2 (Cys) 2 ] site and provided Ni(II)-ligands interatomic distances [18].…”

Nickel Metalloregulators and Chaperones

“…E. coli RcnR coordinates nickel in a six-coordinate octahedral site [70,72,73]. InrS utilizes a different ligand set to coordinate nickel in four-coordinate square planar geometry [79]. Why has nature designed two metalloregulators that carryout similar functions with the same fold but different metal binding geometries and ligand sets?…”

“…His21 (analogous to His3 in RcnR) ( Figure 6) is located on the flexible N-terminal extension and is capable of approaching the Ni(II)-binding site [64]. XAS spectroscopy determined that the nickel site is four-coordinate planar and that His21 is a Ni(II) ligand in InrS [79]. The protein binds Ni(II) using His21, Cys53, His78, and Cys82 forming a Ni(N Im ) 2 (S) 2 complex [79].…”

“…XAS spectroscopy determined that the nickel site is four-coordinate planar and that His21 is a Ni(II) ligand in InrS [79]. The protein binds Ni(II) using His21, Cys53, His78, and Cys82 forming a Ni(N Im ) 2 (S) 2 complex [79]. The protein can also bind 1 Cu(I) per monomer or 1-1.5 Co(II) per monomer [77].…”