PDBsum entry 1git

Gtp-binding protein

PDB id: 1git

Contents

Protein chain

317 a.a. *

Ligands

PO4

GDP

Waters ×9

* Residue conservation analysis

PDB id:

1git

Name:

Gtp-binding protein

Title:

Structure of gtp-binding protein

Structure:

G protein gi alpha 1. Chain: a. Fragment: alpha 1. Engineered: yes. Mutation: yes. Other_details: structure mimics the ternary complex

Source:

Rattus norvegicus. Norway rat. Organism_taxid: 10116. Expressed in: escherichia coli. Expression_system_taxid: 562.

Resolution:

2.60Å R-factor: 0.186 R-free: 0.279

Authors:

A.M.Berghuis,E.Lee,S.R.Sprang

Key ref:

A.M.Berghuis et al. (1996). Structure of the GDP-Pi complex of Gly203-->Ala gialpha1: a mimic of the ternary product complex of galpha-catalyzed GTP hydrolysis. Structure, 4, 1277-1290. PubMed id: 8939752 DOI: 10.1016/S0969-2126(96)00136-0

Date:

16-Oct-96 Release date: 12-Feb-97

Protein chain

P10824  (GNAI1_RAT) -  Guanine nucleotide-binding protein G(i) subunit alpha-1 from Rattus norvegicus

Seq: 354 a.a.

Struc: 317 a.a.*

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

Enzyme reactions

• Enzyme class: E.C.?

DOI no: 10.1016/S0969-2126(96)00136-0

Structure 4:1277-1290 (1996)

PubMed id: 8939752

Structure of the GDP-Pi complex of Gly203-->Ala gialpha1: a mimic of the ternary product complex of galpha-catalyzed GTP hydrolysis.

A.M.Berghuis, E.Lee, A.S.Raw, A.G.Gilman, S.R.Sprang.

ABSTRACT

BACKGROUND: G proteins play a vital role in transmembrane signalling events. In their inactive form G proteins exist as heterotrimers consisting of an alpha subunit, complexed with GDP and a dimer of betagamma subunits. Upon stimulation by receptors, G protein alpha subunits exchange GDP for GTP and dissociate from betagamma . Thus activated, alphasubunits stimulate or inhibit downstream effectors. The duration of the activated state corresponds to the single turnover rate of GTP hydrolysis, which is typically in the range of seconds. In Gialpha1, the Gly203-->Ala mutation reduces the affinity of the substrate for Mg2+, inhibits a key conformational step that occurs upon GTP binding and consequently inhibits the release of betagamma subunits from the GTP complex. The structure of the Gly203-->Ala mutant of Gialpha1 (G203AGialpha1) bound to the slowly hydrolyzing analog of GTP (GTPgammaS) has been determined in order to elucidate the structural changes that take place during hydrolysis. RESULTS: We have determined the three dimensional structure of a Gly203-->Ala mutant of Gialpha1 at 2.6 A resolution. Although crystals were grown in the presence of GTPgammaS and Mg2+, the catalytic site contains a molecule of GDP and a phosphate ion, but no Mg2+. The phosphate ion is bound to a site near that occupied by the gamma-phosphate of GTPgammaS in the activated wild-type alpha subunit. A region of the protein, termed the Switch II helix, twists and bends to adopt a conformation that is radically different from that observed in other Gialpha1 subunit complexes. CONCLUSIONS: Under the conditions of crystallization, the Gly203-->Ala mutation appears to stabilize a conformation that may be similar, although perhaps not identical, to the transient ternary product complex of Gialpha1-catalyzed GTP hydrolysis. The rearrangement of the Switch II helix avoids a potential steric conflict caused by the mutation. However, it appears that dissociation of the gamma-phosphate from the pentacoordinate intermediate also requires a conformational change in Switch II. Thus, a conformational rearrangement of the Switch II helix may be required in Galpha-catalyzed GTP hydrolysis.

Selected figure(s)

Figure 2.
Figure 2. Molecular architecture of G203AG[iα1]. (a) Helical segments are shown as green ribbons and β strands as blue arrows; secondary structural elements near the catalytic site are labeled, GDP and Pi are shown as ball-and-stick models. Switch regions (Sw) are labeled (I–IV) and ‘N’ and ‘C’ mark the locations of residues 34 and 343, respectively. (b) A superposition of the Cα traces of the G[iα1] subunit bound to GDP (blue), G[iα1]bound to GTPγS–Mg^2+ (red), and the GDP–Pi complex of G203AG[iα1] (green). The Cα atoms of residues 40–178 and 220–340 were used to generate the superposition. Figure 2. Molecular architecture of G203AG[iα1]. (a) Helical segments are shown as green ribbons and β strands as blue arrows; secondary structural elements near the catalytic site are labeled, GDP and Pi are shown as ball-and-stick models. Switch regions (Sw) are labeled (I–IV) and ‘N’ and ‘C’ mark the locations of residues 34 and 343, respectively. (b) A superposition of the Cα traces of the G[iα1] subunit bound to GDP (blue), G[iα1]bound to GTPγS–Mg^2+ (red), and the GDP–Pi complex of G203AG[iα1] (green). The Cα atoms of residues 40–178 and 220–340 were used to generate the superposition. (Figure was generated using the program SETOR [[3]55].)

Figure 8.
Figure 8. Conformational states along the reaction pathway (see text). (a) G[iα1]–GTPγS–Mg^2+. (b) Structure of the pentacoordinate intermediate modeled on the structure of the G[iα1]–GDP–AlF[4]^−–Mg^2+ complex. (c) G[iα1]–GDP–Pi complex modeled on the structure of G203A–GDP–Pi. (d) G[iα1]–GDP. Color scheme is the same as that used in Figure 3. Figure 8. Conformational states along the reaction pathway (see text). (a) G[iα1]–GTPγS–Mg^2+. (b) Structure of the pentacoordinate intermediate modeled on the structure of the G[iα1]–GDP–AlF[4]^−–Mg^2+ complex. (c) G[iα1]–GDP–Pi complex modeled on the structure of G203A–GDP–Pi. (d) G[iα1]–GDP. Color scheme is the same as that used in [3]Figure 3. (Figure was generated using the program SETOR [[4]55].)

The above figures are reprinted by permission from Cell Press: Structure (1996, 4, 1277-1290) copyright 1996.

Figures were selected by an automated process.