PDBsum entry 1s2k

Hydrolase

PDB id: 1s2k

Contents

Protein chain

199 a.a. *

Ligands

ALA-ILE-HIS

TYR

Waters ×121

* Residue conservation analysis

PDB id:

1s2k

Name:

Hydrolase

Title:

Structure of scp-b a member of the eqolisin family of peptidases in a complex with a tripeptide ala-ile-his

Structure:

Scytalidopepsin b. Chain: a. Synonym: acid protease b, slb. Engineered: yes. Ala-ile-his tripeptide. Chain: b. Engineered: yes

Source:

Scytalidium lignicola. Organism_taxid: 5539. Expressed in: escherichia coli. Expression_system_taxid: 562. Synthetic: yes. Other_details: tripeptide substrate

Biol. unit:

Dodecamer (from PQS)

Resolution:

2.00Å R-factor: 0.234 R-free: 0.256

Authors:

M.Fujinaga,M.M.Cherney,H.Oyama,K.Oda,M.N.James

Key ref:

M.Fujinaga et al. (2004). The molecular structure and catalytic mechanism of a novel carboxyl peptidase from Scytalidium lignicolum. Proc Natl Acad Sci U S A, 101, 3364-3369. PubMed id: 14993599 DOI: 10.1073/pnas.0400246101

Date:

08-Jan-04 Release date: 27-Apr-04

Protein chain

P15369  (PRTB_SCYLI) -  Scytalidopepsin B from Scytalidium lignicola

Seq: 260 a.a.

Struc: 199 a.a.

Enzyme reactions

• Enzyme class: E.C.3.4.23.32 - scytalidopepsin B.
• Reaction: Hydrolysis of proteins with broad specificity, cleaving 24-Phe-|-Phe-25, but not 15-Leu-|-Tyr-16 and 25-Phe-|-Tyr-26 in the B chain of insulin.

DOI no: 10.1073/pnas.0400246101

Proc Natl Acad Sci U S A 101:3364-3369 (2004)

PubMed id: 14993599

The molecular structure and catalytic mechanism of a novel carboxyl peptidase from Scytalidium lignicolum.

M.Fujinaga, M.M.Cherney, H.Oyama, K.Oda, M.N.James.

ABSTRACT

The molecular structure of the pepstatin-insensitive carboxyl peptidase from Scytalidium lignicolum, formerly known as scytalidopepsin B, was solved by multiple isomorphous replacement phasing methods and refined to an R factor of 0.230 (R(free) = 0.246) at 2.1-A resolution. In addition to the structure of the unbound peptidase, the structure of a product complex of cleaved angiotensin II bound in the active site of the enzyme was also determined. We propose the name scytalidocarboxyl peptidase B (SCP-B) for this enzyme. On the basis of conserved, catalytic residues identified at the active site, we suggest the name Eqolisin for the enzyme family. The previously uninvestigated SCP-B fold is that of a beta-sandwich; each sheet has seven antiparallel strands. A tripeptide product, Ala-Ile-His, bound in the active site of SCP-B has allowed for identification of the catalytic residues and the residues in subsites S1, S2, and S3, which are important for substrate binding. The most likely hydrolytic mechanism involves nucleophilic attack of a general base (Glu-136)-activated water (OH(-)) on the si-face of the scissile peptide carbonylcarbon atom to form a tetrahedral intermediate. Electrophilic assistance and oxyanion stabilization is provided by the side-chain amide of Gln-53. Protonation of the leaving-group nitrogen is accomplished by the general acid function of the protonated carboxyl group of Glu-136.

Selected figure(s)

Figure 4.
Fig. 4. (a) Electron density associated with the product Ala-Ile-His-COO- and the two catalytic residues Gln-53 and Glu-136. The details of the contour levels are as given in Fig. 3a. (b) The environment of the angiotensin II hydrolysis product Ala-Ile-His (orange) is shown. The C terminus of the tripeptide corresponds to a cleavage site in Angiotensin II at the His-Pro bond, and the tripeptide indicates the binding mode of three residues of the N-terminal segment of the substrate. Hydrogen bonds between groups on the enzyme and on the tripeptide are indicated by dotted lines.

Figure 6.
Fig. 6. (a) The proposed catalytic mechanism of SCP-B. The water molecule hydrogen-bonded to both Glu-136 and Gln-53 is the nucleophile. The general base is the carboxylate of Glu-136. The side-chain amide of Gln-53 assists in the nucleophilic attack and stabilizes the tetrahedral intermediate by hydrogen bonding. (b) A model of a substrate Ala-Ile-His-Pro bound in a productive mode in the active site of SCP-B. The nucleophilic attack by the OH- ion (blue sphere) is on the si-face of the scissile peptide. The surface of SCP-B is represented and colored according to the underlying atoms (slate, carbon; blue, nitrogen; red, oxygen).

Figures were selected by an automated process.