{{STRUCTURE_2oxu| PDB=2oxu | SCENE= }}

Matrix Metallopeptidase 12 : 2oxu.

== Introduction ==

2oxu is a human protease. It belongs to the Matrix Metalloproteinase’s family (MMPs). Theses MMPs are secreted as inactive proproteins and then activated after a cleavage by extracellular proteinases.
MMPs are sorted into 17 classes depending on their localization in the cell, and on their substrates. For instance, 2Oxu is a macrophage metalloelastane indexed in MMP12 subfamily. This hydrolysis involves two cofactors: <scene name='User:Julie_Frentzel/Sandbox_1/Zn/1'>zinc</scene> and <scene
name='User:Julie_Frentzel/Sandbox_1/Ca/1'>calcium</scene>. Zn2+ atoms are part of the enzyme whereas Ca2+ atoms are hydrogen bounded to the backbone of the protein.
In healthy cells, 2oxu is implied in tissue remodelling and cell signalling. Its role is to allow the migration of the macrophages in tissues by hydrolysing soluble and insoluble elastin of the extracellular matrix.
Because of its activity, this protein is also involved in metastasis and tumor development; that's why it is considered as an important target for drug therapies.

== From structural data to the enzymatic mecanism ==

Thanks to X-ray crystallography, the structure of 2oxu has been solved with 1,24 Å resolution.
This enzyme consists of <scene name='User:Julie_Frentzel/Sandbox_1/Helixes/1'>four alpha helixes</scene> and <scene name='User:Julie_Frentzel/Sandbox_1/Sheets/1'>five beta sheets</scene> organized like as you can see on the J-mol figure.

The hydrolysis of the Substrate can be divided in four essential events:

1- Substrate's fixation

2- Substrate's cleavage

3- First fragment's release

4- Second fragment's release

The substrate polypeptide sequence which is recognized by the enzyme is the following one : ProGlnGly(206)IleAlaGly(209). It is known to be cleaved between Gly(206) and ILE.
Before the addition of the substrate, the active site is closed.

A video of this hydrolisis is available online [http://www.wiley-vch.de/contents/jc_2002/2006/z603100_s.html]

=== Substrate's fixation ===

The <scene name='User:Julie_Frentzel/Sandbox_1/Active_site/1'>active site of the enzyme</scene> is composed by one '''Glu''' residue near 3 His residues which maintain a <scene name='User:Julie_Frentzel/Sandbox_1/Zinc_et_eau/1'>Zn2+ atom coordinated with three water molecules</scene>. One of them is as well bound to the Glu residue thanks to a hydrogen bond. The second Zn2+ atom is not involved in the active site. At first, the Gly 206 residue of the substrate binds the active site thanks to the Zn2+ atom. When it binds it takes the place of unstable water molecules and establishes stabilizing interactions with the active site thanks to its C terminal part. Then, the Ala 182 residue of the enzyme makes a hydrogen bond with the NH group of the substrate: this allows the substrate to enter the cavity of the catalytic site.
The rest of the protein is stabilized by 4 hydrogen bonds with the amino acid located in the cavity.

=== Substrate's cleavage ===

A proton of one of the three H2O moves to the Glu219 catalytic site's residue.
It creates a hydroxyl group with a high nucleophilic character.
That leads to a nucleophilic attack by zinc-coordinated hydroxyl group, on one of the carbonyl group of the substrate (Gly206).
It leads to what is called a gem-diol intermediate form of the substrate. Two fragments are then sitting in the cavity: IleAlaGly209 and ProGlnGly206.

=== First fragment's release ===

The fragment IleAlaGly is strongly bounded to the enzyme thanks to the bonds that have been created at the fixation's step.
But the fragment ProGlnGly just remains in place thanks to a Gly206 monodentate coordination with the Zn atom.
Then a water molecule binds this Zn atom: it becomes five coordinated, which facilitates the detachment of the ProGlnGly fragment through an associative ligand exchange mechanism.

=== Second fragment's release ===

IleAlaGly remains in the active site but is subjected to a rearrangement.
After the release of ProGlnGly, repulsion between positively charged zinc ion and NH3+ (coming from the peptide's cleavage) increases. It forces IlaAlaGly to enter deeper in the substrate's cavity to make hydrophobic interactions.
The transfer of this fragment prevents Ala208 to continue binding Pro208. The former bound is broken, and the fragment is no more stabilized. Finally the cavity opens and the fragment is released.

== Regulation ==

One of the two Zn2+ ions is involved in the catalytic site to catalyse the reaction. The second Zn2+ atom and the three Ca2+ binds the enzyme to stabilize its 3D strucure.
Ca2+ concentrational change may regulate MMP12 activity.

Some structural modification of 2oxu, can be a way to lower it's enzymatic activity :

*Glu 219 play a very important rôle in the substrate's fixation : if it is mutated and replaced by an non-ionizable residue, it lowers the enzymatic activity.

*If the ionization of NH2 to NH3+ can be prevented, it won't lead to any charge repulsion with Zn2+ : the substrate won't enter the cavity very deeply and so will remain bound to the enzyme, preventing the next binding with the next substrate.

==Research for inhibitors : current stakes ==

MMPs are also thought to play a major role on cell behaviors such as cell proliferation, migration (adhesion/dispersion), differentiation, angiogenesis, apoptosis and host defense.

he MMPs are inhibited by specific endogenous tissue inhibitor of metalloproteinases (TIMPs), which comprise a family of four protease inhibitors: TIMP-1, TIMP-2, TIMP-3 and TIMP-4.
Synthetic inhibitors generally contain a chelating group which binds the catalytic zinc atom at the MMP active site tightly. Common chelating groups include hydroxamates, carboxylates, thiols, and phosphinyls. Hydroxymates are particularly potent inhibitors of MMPs and other zinc-dependent enzymes, due to their bidentate chelation of the zinc atom. Other substitutents of these inhibitors are usually designed to interact with various binding pockets on the MMP of interest, making the inhibitor more or less specific for given MMPs.

== References ==

User:Julie Frentzel/Sandbox 1

2009-11-13T12:33:29Z

Julie Frentzel: /* Regulation */

User:Julie Frentzel/Sandbox 1

2009-11-13T12:33:06Z

Julie Frentzel: /* Regulation */

User:Julie Frentzel/Sandbox 1

2009-11-13T12:29:45Z

Julie Frentzel: /* Introduction */

User:Julie Frentzel/Sandbox 1

2009-11-13T12:27:48Z

Julie Frentzel: /* Structure / function relationship */

User:Julie Frentzel/Sandbox 1

2009-11-06T09:00:43Z

Julie Frentzel: /* Structure / function relationship */