PDBsum entry 2vui

DNA-binding

PDB id: 2vui

Contents

Protein chain

137 a.a. *

Ligands

BEF

Metals

_MG ×2

* Residue conservation analysis

PDB id:

2vui

Name:

DNA-binding

Title:

Crystal structure of the hupr receiver domain in inhibitory phospho- state

Structure:

Hydrogenase transcriptional regulatory protein hupr1. Chain: b. Fragment: receiver domain, residues 5-140. Synonym: hupr. Engineered: yes

Source:

Rhodobacter capsulatus. Organism_taxid: 1061. Expressed in: escherichia coli. Expression_system_taxid: 469008.

Resolution:

2.90Å R-factor: 0.261 R-free: 0.320

Authors:

K.M.Davies,E.D.Lowe,C.Venien-Bryan,L.N.Johnson

Key ref:

K.M.Davies et al. (2009). The HupR receiver domain crystal structure in its nonphospho and inhibitory phospho states. J Mol Biol, 385, 51-64. PubMed id: 18977359 DOI: 10.1016/j.jmb.2008.10.027

Date:

26-May-08 Release date: 11-Nov-08

Protein chain

P26408  (HUPR_RHOCA) -  Hydrogenase transcriptional regulatory protein HupR1 from Rhodobacter capsulatus

Seq: 492 a.a.

Struc: 137 a.a.*

* PDB and UniProt seqs differ at 1 residue position (black cross)

DOI no: 10.1016/j.jmb.2008.10.027

J Mol Biol 385:51-64 (2009)

PubMed id: 18977359

The HupR receiver domain crystal structure in its nonphospho and inhibitory phospho states.

K.M.Davies, E.D.Lowe, C.Vénien-Bryan, L.N.Johnson.

ABSTRACT

Hydrogen uptake protein regulator (HupR) is a member of the nitrogen regulatory protein C (NtrC) family of response regulators. These proteins activate transcription by RNA polymerase (RNAP) in response to a change in environment. This change is detected through the phosphorylation of their receiver domain as part of a two-component signalling pathway. HupR is an unusual member of this family as it activates transcription when unphosphorylated, and transcription is inhibited by phosphorylation. Also, HupR activates transcription through the more general sigma(70) transcription initiation factor, which does not require activation by ATPase, in contrast to other NtrC family members that utilise sigma(54). Hence, its mode of action is expected to be different from those of the more conventional NtrC family members. We have determined the structures of the unphosphorylated N-terminal receiver domain of wild-type HupR, the mutant HupR(D55E)(N) (which cannot be phosphorylated and down-regulated), and HupR in the presence of the phosphorylation mimic BeF(3)(-). The structures show a typical response regulator fold organised as a dimer whose interface involves alpha4-beta5-alpha5 elements. The interactions across the interface are slightly different between apo and phospho mimics, and these reflect a rearrangement of key conserved residues around the active site aspartate that have been implicated in domain activation in other receiver domain proteins. We also show that the wild-type HupR receiver domain forms a weak dimer in solution, which is strengthened in the presence of the phosphorylation mimic BeF(3)(-). The results indicate many features similar to those that have been observed in other systems, including NtrC (where phosphorylation is activatory), and indicate that recognition properties, which allow HupR to be active in the absence of phosphorylation, lie in the transmission of phosphorylation signals through the linker region to the other domains of the protein.

Selected figure(s)

Figure 1.
Fig. 1. The structure of HupR[WT]^N. (a) Cartoon representation of HupR[WT]^N. The site of phosphorylation (Asp55) is illustrated as green sticks. The position of the conserved residues involved in activation, Thr83 and Phe103, is illustrated as yellow and blue sticks, respectively. Secondary structure elements are labelled as follows: β1, residues 7–12; β2, residues 30–35; β3, residues 48–55; β4, residues 78–83; β5, residues 101–105; α1, residues 15–28; α2, residues 36–46; α3, residues 63–73; α4, residues 89–98; α5, residues 109–141. (b) Magnesium binding at the active site of HupR[WT]^N. Mg^2 + (pink sphere) and H[2]O (blue sphere). No unexplained density is observed around Asp55; thus, HupR[WT]^N is unphosphorylated. Blue density corresponds to the 2F[o] − F[c] map contoured at 1σ, and pink density corresponds to the F[o] − F[c] map contoured at 5σ, with Mg^2 + excluded from the phasing. (c) Cartoon representation of a HupR[WT]^N dimer viewed along the crystallographic 2-fold axis. The dimer interface occurs between β5 and α5 of neighbouring monomers. The phosphorylatable aspartate Asp55 is shown as green sticks, and the conserved aromatic residue Phe103 is shown as blue sticks. Figures were created with PyMOL™ (DeLano Scientific LLC).

Figure 2.
Fig. 2. The structure of HupR[D55E]^N. (a) Overlay of HupR[WT]^N and HupR[D55E]^N[.] The two structures were aligned according to residues 6–81. Thr83 and Phe103 are illustrated as sticks: Thr83-HupR[WT]^N (yellow), Thr83-HupR[D55E]^N (orange), Phe103-HupR[WT]^N (cyan), and Phe103-HupR[D55E]^N (light blue). (b) Active site of HupR[WT]^N and HupR[D55E]^N showing the different conformations of Thr83 and Phe103. In HupR[WT]^N, Thr83 points away from Asp55, and Phe103 points out of the monomer. In HupR[D55E]^N, Thr83 forms a hydrogen bond with Glu55, and Phe103 points into the monomer. (c) Dimer interface of HupR[WT]^N and HupR[D55E]^N showing the different interactions that occur across the interface. Hydrophobic residues: salmon sticks; hydrogen bonds: black dashes.

The above figures are reprinted by permission from Elsevier: J Mol Biol (2009, 385, 51-64) copyright 2009.

Figures were selected by the author.