PDBsum entry 1obk

Amidase

PDB id: 1obk

Contents

Protein chains

413 a.a. *

Waters ×925

* Residue conservation analysis

PDB id:

1obk

Name:

Amidase

Title:

Crystal structure of the r158q mutant of malonamidase e2 from bradyrhizobium japonicum

Structure:

Malonamidase e2. Chain: a, b. Engineered: yes. Mutation: yes. Other_details: cis peptide bond between glycine 130 and serine 131

Source:

Bradyrhizobium japonicum. Organism_taxid: 375. Expressed in: escherichia coli. Expression_system_taxid: 469008.

Biol. unit:

Dimer (from PDB file)

Resolution:

2.20Å R-factor: 0.177 R-free: 0.251

Authors:

S.Shin,B.-H.Oh

Key ref:

S.Shin et al. (2003). Characterization of a novel Ser-cisSer-Lys catalytic triad in comparison with the classical Ser-His-Asp triad. J Biol Chem, 278, 24937-24943. PubMed id: 12711609 DOI: 10.1074/jbc.M302156200

Date:

31-Jan-03 Release date: 13-Feb-04

Protein chains

Q9ZIV5  (Q9ZIV5_BRAJP) -  Malonamidase E2 from Bradyrhizobium japonicum

Seq: 414 a.a.

Struc: 413 a.a.*

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

DOI no: 10.1074/jbc.M302156200

J Biol Chem 278:24937-24943 (2003)

PubMed id: 12711609

Characterization of a novel Ser-cisSer-Lys catalytic triad in comparison with the classical Ser-His-Asp triad.

S.Shin, Y.S.Yun, H.M.Koo, Y.S.Kim, K.Y.Choi, B.H.Oh.

ABSTRACT

Amidase signature family enzymes, which are widespread in nature, contain a newly identified Ser-cisSer-Lys catalytic triad in which the peptide bond between Ser131 and the preceding residue Gly130 is in a cis configuration. In order to characterize the property of the novel triad, we have determined the structures of five mutant malonamidase E2 enzymes that contain a Cys-cisSer-Lys, Ser-cisAla-Lys, or Ser-cisSer-Ala triad or a substitution of Gly130 with alanine. Cysteine cannot replace the role of Ser155 due to a hyper-reactivity of the residue, which results in the modification of the cysteine to cysteinyl sulfinic acid, most likely inside the expression host cells. The lysine residue plays a structural as well as a catalytic role, since the substitution of the residue with alanine disrupts the active site structure completely. The two observations are in sharp contrast with the consequences of the corresponding substitutions in the classical Ser-His-Asp triad. Structural data on the mutant containing the Ser-cisAla-Lys triad convincingly suggest that Ser131 plays an analogous catalytic role as the histidine of the Ser-His-Asp triad. The unusual cis configuration of Ser131 appears essential for the precise contacts of this residue with the other triad residues, as indicated by the near invariance of the preceding glycine residue (Gly130), structural data on the G130A mutant, and by a modeling experiment. The data provide a deep understanding of the role of each residue of the new triad at the atomic level and demonstrate that the new triad is a catalytic device distinctively different from the classical triad or its variants.

Selected figure(s)

Figure 5.
FIG. 5. G130A mutation fails to maintain a structured loop. A stereo view of the 2F[o]-F[c] map (2.2 Å, 1 ) of the G130A mutant is shown for the active site. Although Ala^130 was included in the final refinement and map calculation, electron density for this residue is only partly visible.

Figure 6.
FIG. 6. K62A mutation disrupts the active site. a, 2F[o]-F[c] electron density map for the active site of the K62A mutant. The map was calculated to 2.0 Å and contoured at 1.5 . Two tightly bound water molecules, Wat942 and Wat941, indicated as Wat1 and Wat2, are absent in the wild-type MAE2 structure. The hydrogen bonds present only in the mutant structure are shown in dotted lines. b, superposition of the active sites. The catalytic components of the wild-type (in coral) and K62A mutant (in blue) enzymes are superimposed. The circles indicate the positions of the Ser155 in the two structures. Unlike the oxyanion hole-containing segment, the cisSer131-containing loop undergoes a minor conformational change. Figs. 1, 2, 3, 4, 5, 6 were prepared using the program BobScript and rendered using Raster3D.

The above figures are reprinted by permission from the ASBMB: J Biol Chem (2003, 278, 24937-24943) copyright 2003.

Figures were selected by an automated process.