PDBsum entry 2zxc

Hydrolase

PDB id: 2zxc

Contents

Protein chains

643 a.a. *

Ligands

2ED ×2

FMT ×8

DMS ×2

Metals

_MG ×2

_ZN ×2

Waters ×939

* Residue conservation analysis

PDB id:

2zxc

Name:

Hydrolase

Title:

Ceramidase complexed with c2

Structure:

Neutral ceramidase. Chain: a, b. Synonym: ncdase, acylsphingosine deacylase, n-acylsphingosine amidohydrolase. Engineered: yes

Source:

Pseudomonas aeruginosa. Organism_taxid: 287. Expressed in: escherichia coli. Expression_system_taxid: 562

Resolution:

2.20Å R-factor: 0.194 R-free: 0.228

Authors:

H.Okano,T.Inoue,N.Okino,Y.Kakuta,H.Matsumura,M.Ito

Key ref:

T.Inoue et al. (2009). Mechanistic insights into the hydrolysis and synthesis of ceramide by neutral ceramidase. J Biol Chem, 284, 9566-9577. PubMed id: 19088069 DOI: 10.1074/jbc.M808232200

Date:

22-Dec-08 Release date: 03-Feb-09

Protein chains

Q9I596  (NCASE_PSEAE) -  Neutral ceramidase from Pseudomonas aeruginosa (strain ATCC 15692 / DSM 22644 / CIP 104116 / JCM 14847 / LMG 12228 / 1C / PRS 101 / PAO1)

Seq: 670 a.a.

Struc: 643 a.a.*

* PDB and UniProt seqs differ at 3 residue positions (black crosses)

Enzyme reactions

• Enzyme class: E.C.3.5.1.23 - ceramidase.
• Reaction: an N-acylsphing-4-enine + H2O = sphing-4-enine + a fatty acid

N-acylsphing-4-enine + H2O = sphing-4-enine (Bound ligand (Het Group name = FMT) matches with 75.00% similarity) + fatty acid (Bound ligand (Het Group name = 2ED) matches with 41.67% similarity)

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

Added reference

DOI no: 10.1074/jbc.M808232200

J Biol Chem 284:9566-9577 (2009)

PubMed id: 19088069

Mechanistic insights into the hydrolysis and synthesis of ceramide by neutral ceramidase.

T.Inoue, N.Okino, Y.Kakuta, A.Hijikata, H.Okano, H.M.Goda, M.Tani, N.Sueyoshi, K.Kambayashi, H.Matsumura, Y.Kai, M.Ito.

ABSTRACT

Ceramidase (CDase; EC 3.5.1.23) hydrolyzes ceramide to generate sphingosine and fatty acid. The enzyme plays a regulatory role in a variety of physiological events in eukaryotes and also functions as an exotoxin in particular bacteria. The crystal structures of neutral CDase from Pseudomonas aeruginosa (PaCD) in the C2-ceramide-bound and -unbound forms were determined at 2.2 and 1.4 A resolutions, respectively. PaCD consists of two domains, and the Zn(2+)- and Mg(2+)/Ca(2+)-binding sites are found within the center of the N-terminal domain and the interface between the domains, respectively. The structural comparison between the C2-ceramide-bound and unbound forms revealed an open-closed conformational change occurring to loop I upon binding of C2-ceramide. In the closed state, this loop sits above the Zn(2+) coordination site and over the opening to the substrate binding site. Mutational analyses of residues surrounding the Zn(2+) of PaCD and rat neutral CDase revealed that the cleavage or creation of the N-acyl linkage of ceramide follows a similar mechanism as observed for the Zn(2+)-dependent carboxypeptidases. The results provide an understanding of the molecular mechanism of hydrolysis and synthesis of ceramide by the enzyme. Furthermore, insights into the actions of PaCD and eukaryotic neutral CDases as an exotoxin and mediators of sphingolipid signaling are also revealed, respectively.

Selected figure(s)

Figure 3.
C2-Cer bound in the putative active site of PaCD and a proposed reactionmechanism.A, the structural comparison between substrate-unbound and C2 Cer-bound forms of PaCD. The substrate-unbound form is shown in red, whereas the C2 Cer-bound form is shown in green. B, the binding model of C2-Cer is shown as a stick representation with oxygen atoms in red, nitrogen atoms in blue, and carbon atoms in yellow. The electron density map was calculated at the 1.0 σ level. C, schematic drawing of the binding model of C2-Cer. The hydrogen bonds and the metal coordinate ion structure was shown by dotted lines. D, a proposed reaction mechanism of PaCD based on the structural data.

Figure 5.
The proposed conformations of nCDase acting on membrane-bound Cer (B and C) and free Cer (A and D). nCDase is shown as a ribbon model. The type II integral membrane (A and B) and soluble (C and D) forms of nCDase are shown.

The above figures are reprinted by permission from the ASBMB: J Biol Chem (2009, 284, 9566-9577) copyright 2009.

Figures were selected by an automated process.