PDBsum entry: 1c8t

Hydrolase/hydrolase inhibitor

PDB id: 1c8t

Contents

Protein chains

167 a.a. *

Ligands

TR1 ×2

Metals

_ZN ×4

_CA ×6

Waters ×71

* Residue conservation analysis

PDB id:

1c8t

Name:

Hydrolase/hydrolase inhibitor

Title:

Human stromelysin-1 (e202q) catalytic domain complexed with ro-26-2812

Structure:

Stromelysin-1. Chain: a, b. Fragment: catalytic domain. Engineered: yes. Mutation: yes

Source:

Homo sapiens. Human. Organism_taxid: 9606. Cell_line: gigival fibroblasts. Expressed in: escherichia coli. Expression_system_taxid: 562.

Resolution:

2.60Å R-factor: 0.220

Authors:

D.L.Steele,O.El-Kabbani,P.Dunten,R.L.Crowther

Key ref:

D.L.Steele et al. (2000). Expression, characterization and structure determination of an active site mutant (Glu202-Gln) of mini-stromelysin-1. Protein Eng, 13, 397-405. PubMed id: 10877850 DOI: 10.1093/protein/13.6.397

Date:

29-Jul-99 Release date: 19-Jul-00

Protein chains

P08254  (MMP3_HUMAN) -  Stromelysin-1

Seq: 477 a.a.

Struc: 167 a.a.*

* PDB and UniProt seqs differ at 2 residue positions (black crosses)

Enzyme reactions

• Enzyme class: E.C.3.4.24.17 - Stromelysin 1.
• Reaction: Preferential cleavage where P1', P2' and P3' are hydrophobic residues.
• Cofactor: Calcium; Zinc

 Gene Ontology (GO) functional annotation 

• Cellular component: extracellular matrix (1 term)
• Biological process: proteolysis (1 term)
• Biochemical function: metallopeptidase activity (3 terms)

DOI no: 10.1093/protein/13.6.397

Protein Eng 13:397-405 (2000)

PubMed id: 10877850

Expression, characterization and structure determination of an active site mutant (Glu202-Gln) of mini-stromelysin-1.

D.L.Steele, O.El-Kabbani, P.Dunten, L.J.Windsor, R.U.Kammlott, R.L.Crowther, C.Michoud, J.A.Engler, J.J.Birktoft.

ABSTRACT

Human stromelysin-1 is a member of the matrix metalloproteinase (MMP) family of enzymes. The active site glutamic acid of the MMPs is conserved throughout the family and plays a pivotal role in the catalytic mechanism. The structural and functional consequences of a glutamate to glutamine substitution in the active site of stromelysin-1 were investigated in this study. In contrast to the wild-type enzyme, the glutamine-substituted mutant was not active in a zymogram assay where gelatin was the substrate, was not activated by organomercurials and showed no activity against a peptide substrate. The glutamine-substituted mutant did, however, bind to TIMP-1, the tissue inhibitor of metalloproteinases, after cleavage of the propeptide with trypsin. A second construct containing the glutamine substitution but lacking the propeptide was also inactive in the proteolysis assays and capable of TIMP-1 binding. X-ray structures of the wild-type and mutant proteins complexed with the propeptide-based inhibitor Ro-26-2812 were solved and in both structures the inhibitor binds in an orientation the reverse of that of the propeptide in the pro-form of the enzyme. The inhibitor makes no specific interactions with the active site glutamate and a comparison of the wild-type and mutant structures revealed no major structural changes resulting from the glutamate to glutamine substitution.