PDBsum entry 2dbu

Transferase

PDB id: 2dbu

Contents

Protein chains

359 a.a. *

190 a.a. *

Waters ×605

* Residue conservation analysis

PDB id:

2dbu

Name:

Transferase

Title:

Crystal structure of gamma-glutamyltranspeptidase from escherichia coli

Structure:

Gamma-glutamyltranspeptidase. Chain: a, c. Fragment: large subunit. Engineered: yes. Gamma-glutamyltranspeptidase. Chain: b, d. Fragment: small subunit. Engineered: yes

Source:

Escherichia coli k12. Organism_taxid: 83333. Strain: k-12. Gene: ggt. Expressed in: escherichia coli. Expression_system_taxid: 562.

Biol. unit:

Dimer (from PQS)

Resolution:

1.95Å R-factor: 0.207 R-free: 0.231

Authors:

T.Okada,K.Wada,K.Fukuyama

Key ref:

T.Okada et al. (2006). Crystal structures of gamma-glutamyltranspeptidase from Escherichia coli, a key enzyme in glutathione metabolism, and its reaction intermediate. Proc Natl Acad Sci U S A, 103, 6471-6476. PubMed id: 16618936 DOI: 10.1073/pnas.0511020103

Date:

16-Dec-05 Release date: 18-Apr-06

Protein chains

P18956  (GGT_ECOLI) -  Glutathione hydrolase proenzyme from Escherichia coli (strain K12)

Seq: 580 a.a.

Struc: 359 a.a.

Protein chains

P18956  (GGT_ECOLI) -  Glutathione hydrolase proenzyme from Escherichia coli (strain K12)

Seq: 580 a.a.

Struc: 190 a.a.

Enzyme reactions

• Enzyme class 2: Chains A, B, C, D: E.C.2.3.2.2 - gamma-glutamyltransferase.
• Reaction: an N-terminal (5-L-glutamyl)-[peptide] + an alpha-amino acid = 5-L- glutamyl amino acid + an N-terminal L-alpha-aminoacyl-[peptide]
• Enzyme class 3: Chains A, B, C, D: E.C.3.4.19.13 - glutathione gamma-glutamate hydrolase.
• Reaction:
  1. glutathione + H2O = L-cysteinylglycine + L-glutamate
  2. an S-substituted glutathione + H2O = an S-substituted L-cysteinylglycine + L-glutamate

Note, where more than one E.C. class is given (as above), each may correspond to a different protein domain or, in the case of polyprotein precursors, to a different mature protein.

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

reference

DOI no: 10.1073/pnas.0511020103

Proc Natl Acad Sci U S A 103:6471-6476 (2006)

PubMed id: 16618936

Crystal structures of gamma-glutamyltranspeptidase from Escherichia coli, a key enzyme in glutathione metabolism, and its reaction intermediate.

T.Okada, H.Suzuki, K.Wada, H.Kumagai, K.Fukuyama.

ABSTRACT

Gamma-glutamyltranspeptidase (GGT) is a heterodimic enzyme that is generated from the precursor protein through posttranslational processing and catalyzes the hydrolysis of gamma-glutamyl bonds in gamma-glutamyl compounds such as glutathione and/or the transfer of the gamma-glutamyl group to other amino acids and peptides. We have determined the crystal structure of GGT from Escherichia coli K-12 at 1.95 A resolution. GGT has a stacked alphabetabetaalpha fold comprising the large and small subunits, similar to the folds seen in members of the N-terminal nucleophile hydrolase superfamily. The active site Thr-391, the N-terminal residue of the small subunit, is located in the groove, from which the pocket for gamma-glutamyl moiety binding follows. We have further determined the structure of the gamma-glutamyl-enzyme intermediate trapped by flash cooling the GGT crystal soaked in glutathione solution and the structure of GGT in complex with l-glutamate. These structures revealed how the gamma-glutamyl moiety and l-glutamate are recognized by the enzyme. A water molecule was seen on the carbonyl carbon of the gamma-glutamyl-Thr-391 Ogamma bond in the intermediate that is to be hydrolyzed. Notably the residues essential for GGT activity (Arg-114, Asp-433, Ser-462, and Ser-463 in E. coli GGT) shown by site-directed mutagenesis of human GGT are all involved in the binding of the gamma-glutamyl moiety. The structure of E. coli GGT presented here, together with sequence alignment of GGTs, may be applicable to interpret the biochemical and genetic data of other GGTs.

Selected figure(s)

Figure 1.
Fig. 1. Structure of E. coli GGT. (a) Ribbon drawing of the GGT heterodimer. The L subunit is colored blue, and the S subunit is colored green. (b) Ribbon drawing of the L subunit. (c) Ribbon drawing of the S subunit. -Helices and -strands are labeled. In each of the L and S subunits, the N terminus is blue and the C terminus is red, with intermediate colors following the distance in the sequence from the N terminus. The N-terminal residue of the S subunit (Thr-391) is shown with a stick model. (d) A topology diagram of E. coli GGT. Circle, triangle, and square indicate -helix, -strand, and insertion not conserved among Ntn-hydrolases, respectively. The secondary structures were defined with DSSP (19). The figures were prepared with PYMOL (20) and TOPS (21).

Figure 2.
Fig. 2. The structure of the substrate binding pocket of E. coli GGT. (a) Surface drawing of substrate binding pocket. The stick model of the -glutamyl moiety, nucleophile (Thr-391), and residues forming the wall (Asn-411 and Tyr-444) are shown in blue, green, and yellow, respectively. Green dots represent the groove in which the peptide of the precursor protein is assumed to be present. The hydrogen bond between Asn-411 O and Tyr-444 O is shown as a dashed line. The ribbon model shown in yellow represents residues Pro-438–Gly-449, which are absent in B. subtilis GGT. (b) The (F[o] – F[c]) omit map contoured at the 3 level for GGT- G. The omit map was generated by omitting the -glutamyl moiety, Thr-391, and a water molecule (labeled W2) from the model. Ball-and-stick models of -glutamyl–enzyme complex are overlaid on the map. The residues involved in substrate binding and enzyme reaction are shown in the model. For the clarity, the side chains of Gln-89, Leu-410, and Thr-412 are omitted from the model. Water molecules involved in substrate binding and the catalytic reaction are labeled (W1 W3). The hydrogen bonds are shown as dashed lines. (c) The (F[o] – F[c]) omit map for GGT-Glu prepared as for GGT- G. The view direction is rotated by 40° around the vertical axis relative to b. The figures were prepared with PYMOL (20).

Figures were selected by an automated process.