PDBsum entry 2j9d

Membrane transport

PDB id: 2j9d

Contents

Protein chains

(+ 0 more) 102 a.a. *

(+ 0 more) 116 a.a. *

Ligands

ACT ×5

ADP ×4

AMP

Metals

_CL

Waters ×694

* Residue conservation analysis

PDB id:

2j9d

Name:

Membrane transport

Title:

Structure of glnk1 with bound effectors indicates regulatory mechanism for ammonia uptake

Structure:

Hypothetical nitrogen regulatory pii-like protein mj0059. Chain: a, b, c, d, f, g, h, i, j, k, l. Synonym: glnk1. Engineered: yes. Hypothetical nitrogen regulatory pii-like protein mj0059. Chain: e. Synonym: glnk1. Engineered: yes

Source:

Methanococcus jannaschii. Organism_taxid: 2190. Strain: amjft37. Atcc: 625482. Expressed in: escherichia coli. Expression_system_taxid: 469008.

Resolution:

2.10Å R-factor: 0.210 R-free: 0.265

Authors:

O.Yildiz,C.Kalthoff,S.Raunser,W.Kuehlbrandt

Key ref:

O.Yildiz et al. (2007). Structure of GlnK1 with bound effectors indicates regulatory mechanism for ammonia uptake. EMBO J, 26, 589-599. PubMed id: 17203075 DOI: 10.1038/sj.emboj.7601492

Date:

07-Nov-06 Release date: 16-Jan-07

Protein chains

Q60381  (Y059_METJA) -  Nitrogen regulatory protein GlnK1 from Methanocaldococcus jannaschii (strain ATCC 43067 / DSM 2661 / JAL-1 / JCM 10045 / NBRC 100440)

Seq: 112 a.a.

Struc: 102 a.a.

Protein chains

Q60381  (Y059_METJA) -  Nitrogen regulatory protein GlnK1 from Methanocaldococcus jannaschii (strain ATCC 43067 / DSM 2661 / JAL-1 / JCM 10045 / NBRC 100440)

Seq: 112 a.a.

Struc: 116 a.a.

DOI no: 10.1038/sj.emboj.7601492

EMBO J 26:589-599 (2007)

PubMed id: 17203075

Structure of GlnK1 with bound effectors indicates regulatory mechanism for ammonia uptake.

O.Yildiz, C.Kalthoff, S.Raunser, W.Kühlbrandt.

ABSTRACT

A binary complex of the ammonia channel Amt1 from Methanococcus jannaschii and its cognate P(II) signalling protein GlnK1 has been produced and characterized. Complex formation is prevented specifically by the effector molecules Mg-ATP and 2-ketoglutarate. Single-particle electron microscopy of the complex shows that GlnK1 binds on the cytoplasmic side of Amt1. Three high-resolution X-ray structures of GlnK1 indicate that the functionally important T-loop has an extended, flexible conformation in the absence of Mg-ATP, but assumes a compact, tightly folded conformation upon Mg-ATP binding, which in turn creates a 2-ketoglutarate-binding site. We propose a regulatory mechanism by which nitrogen uptake is controlled by the binding of both effector molecules to GlnK1. At normal effector levels, a 2-ketoglutarate molecule binding at the apex of the compact T-loop would prevent complex formation, ensuring uninhibited ammonia uptake. At low levels of Mg-ATP, the extended loops would seal the ammonia channels in the complex. Binding of both effector molecules to P(II) signalling proteins may thus represent an effective feedback mechanism for regulating ammonium uptake through the membrane.

Selected figure(s)

Figure 4.
Figure 4 X-ray structures of GlnK1 (A) in the absence of added nucleotide at 2.1 Å; (B) with bound Mg-ATP at 1.2 Å; (C) with Mg-ATP and 2-KG at 1.6 Å resolution. On the left is the trimer with the Amt1 interaction surface facing up. Individual protein monomers are drawn as blue, green and gray ribbon diagrams. Bound cofactors are shown as ball-and-stick models. In (A), one out of three monomers binds ADP; in (B), all three monomers bind Mg-ATP; in (C), all three monomers bind ATP, but only the blue monomer binds Mg^2+ as well, plus a single molecule of 2-KG. The center of the trimer holds either an acetate (A) or a chloride ion (yellow sphere). The corresponding monomer side views are shown on the right. (A) Superposition of six monomers with resolved T-loops in the extended conformation (Supplementary Figure 4). (B) Superposition of three monomers of one Mg-ATP-binding trimer, with T-loops in the compact conformation. Color coding as in Supplementary Figures 3 and 4. (C) Single monomer binding Mg-ATP and 2-KG.

Figure 5.
Figure 5 Stereo diagrams showing details of the electron density of T-loop residues and the nucleotide-binding pocket at the interface between two adjacent GlnK1 monomers (blue and green). (A) Without added Mg-ATP, occasional sites are occupied by ADP (red) or AMP from the expressing cells. The T-loop is in an extended conformation, with arginines 45, 47 and 49 and Tyr51at the tip. (B) Mg-ATP (red) fixes the T-loop in the compact conformation through main-chain interactions with the ATP -phosphate and hydrogen bonds with Mg-coordinated water molecules, positioning Glu44 to form a salt bridge with Lys58. (C) By fixing the T-loop in its compact conformation, Mg-ATP (red) creates a binding site for 2-KG (yellow) on the far side of Tyr51. The density above the keto oxygen of 2-KG is due to an ordered water molecule.

The above figures are reprinted from an Open Access publication published by Macmillan Publishers Ltd: EMBO J (2007, 26, 589-599) copyright 2007.

Figures were selected by an automated process.