PDBsum entry: 2vf4

Transferase

PDB id: 2vf4

Contents

Protein chain

359 a.a. *

Waters ×11

* Residue conservation analysis

PDB id:

2vf4

Name:

Transferase

Title:

E. Coli glucosamine-6-p synthase

Structure:

Glucosamine--fructose-6-phosphate aminotransferas chain: x. Fragment: residues 2-609. Synonym: hexosephosphate aminotransferase, d-fructose-6-pho amidotransferase, gfat, l-glutamine-d- fructose-6-phosphateamidotransferase, glucosamine-6-phosph synthase, glucosamine-6-p synthase. Engineered: yes

Source:

Escherichia coli. Organism_taxid: 562. Expressed in: escherichia coli. Expression_system_taxid: 562.

Resolution:

2.95Å R-factor: 0.220 R-free: 0.257

Authors:

S.Mouilleron,B.Golineli-Pimpaneau

Key ref:

S.Mouilleron et al. (2008). Ordering of C-terminal loop and glutaminase domains of glucosamine-6-phosphate synthase promotes sugar ring opening and formation of the ammonia channel. J Mol Biol, 377, 1174-1185. PubMed id: 18295797 DOI: 10.1016/j.jmb.2008.01.077

Date:

30-Oct-07 Release date: 04-Mar-08

Protein chain

P17169  (GLMS_ECOLI) -  Glucosamine--fructose-6-phosphate aminotransferase [isomerizing]

Seq: 609 a.a.

Struc: 359 a.a.

Enzyme reactions

• Enzyme class: E.C.2.6.1.16 - Glutamine--fructose-6-phosphate transaminase (isomerizing).
• Pathway: UDP-N-acetylglucosamine Biosynthesis
• Reaction: L-glutamine + D-fructose 6-phosphate = L-glutamate + D-glucosamine 6-phosphate

Molecule diagrams generated from .mol files obtained from the KEGG ftp site

 Gene Ontology (GO) functional annotation 

• Cellular component: cytoplasm (2 terms)
• Biological process: metabolic process (5 terms)
• Biochemical function: protein binding (5 terms)

reference

DOI no: 10.1016/j.jmb.2008.01.077

J Mol Biol 377:1174-1185 (2008)

PubMed id: 18295797

Ordering of C-terminal loop and glutaminase domains of glucosamine-6-phosphate synthase promotes sugar ring opening and formation of the ammonia channel.

S.Mouilleron, M.A.Badet-Denisot, B.Golinelli-Pimpaneau.

ABSTRACT

Glucosamine-6-phosphate synthase (GlmS) channels ammonia from glutamine at the glutaminase site to fructose 6-phosphate (Fru6P) at the synthase site. Escherichia coli GlmS is composed of two C-terminal synthase domains that form the dimer interface and two N-terminal glutaminase domains at its periphery. We report the crystal structures of GlmS alone and in complex with the glucosamine-6-phosphate product at 2.95 A and 2.9 A resolution, respectively. Surprisingly, although the whole protein is present in this crystal form, no electron density for the glutaminase domain was observed, indicating its mobility. Comparison of the two structures with that of the previously reported GlmS-Fru6P complex shows that, upon sugar binding, the C-terminal loop, which forms the major part of the channel walls, becomes ordered and covers the synthase site. The ordering of the glutaminase domains likely follows Fru6P binding by the anchoring of Trp74, which acts as the gate of the channel, on the closed C-terminal loop. This is accompanied by a major conformational change of the side chain of Lys503# of the neighboring synthase domain that strengthens the interactions of the synthase domain with the C-terminal loop and completely shields the synthase site. The concomitant conformational change of the Lys503#-Gly505# tripeptide places catalytic His504# in the proper position to open the sugar and buries the linear sugar, which is now in the vicinity of the catalytic groups involved in the sugar isomerization reaction. Together with the previously reported structures of GlmS in complex with Fru6P or glucose 6-phosphate and a glutamine analogue, the new structures reveal the structural changes occurring during the whole catalytic cycle.

Selected figure(s)

Figure 3.
Fig. 3. In the absence of sugar, the C-tail is disordered and the synthase site is accessible to solvent. (a) 2 F[o] - F[cal] electron density map contoured at 1 σ shows no density for residues 602–608 of the C-tail in the GlmS structure, which indicates its disorder. Two synthase domains (stick models in yellow and orange) belonging to neighboring asymmetric units form an extensive interface. In the GlmS–GlcN6P structure (synthase domains in pink and magenta coils), the C-tail covers the GlcN6P product (sphere model) in the synthase site. (b) Synthase site of the GlmS–GlcN6P structure showing the GlcN6P ligand and residues distant less than 4 Å. A 2 F[o] – F[cal] electron density map contoured at 1 σ is superimposed on the model. (c) General view of the sugar-binding site. The synthase active site is formed at the interface of two synthase domains belonging to different monomers (in dark blue and light blue). When both the synthase and glutaminase domains are ordered as in the GlmS–Fru6P structure^4 (glutaminase domain in cyan), the C-tail (in red) is sandwiched between one synthase domain and one glutaminase domain of the same monomer. The sugar-binding site is made by the backbones of the His-loop (in green) and C-tail. The Trp74 indole group from the glutaminase domain also participates in shielding the synthase site from solvent. The opening of the synthase site may result from a movement of the C-tail, the His-loop and/or the glutaminase domain.

Figure 5.
Fig. 5. Step-by-step formation of the ammonia channel. The accessible surface in the different structures calculated with PyMol (http://pymol.sourceforge.net/) and a probe of 1.3 Å radius is represented following the same color scheme as Fig. 4 and is superposed to the accessible surface of the channel in the GlmS–Glc6P–DON structure, represented as a gray mesh surface. The positions of Glc6P and DON as observed in the GlmS–Glc6P–DON structure are indicated in stick representation to locate the synthase and glutaminase sites in all structures. (a) The GlmS structure. The glutaminase domain and the C-tail are not ordered and the synthase site is open to solvent. The channel is not formed because only the His-loop forms one of its rims. (b) The GlmS–GlcN6P structure. In the presence of cyclic sugars, the C-tail is in a relaxed conformation and Lys503^# participates in shielding the synthase site from solvent in the absence of ordered glutaminase domains. (c) The GlmS–Fru6P structure. Because of the ordering of the glutaminase domains, the channel is almost formed but not continuous because the indole group of Trp74 is inserted between the two observed adjacent cavities. (d) The Glms–Glc6P–DON structure. The rotation of the Trp74 indole group opens the channel, which is fully functional.

The above figures are reprinted by permission from Elsevier: J Mol Biol (2008, 377, 1174-1185) copyright 2008.

Figures were selected by an automated process.