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Flashcards in MMP Deck (13):
1

Matrix Metalloproteinases

The matrix metalloproteinases (MMPs) are a family of ____ metalloproteinases - these are ___+- and ____-dependent ____ that are active at ____ pH

MMPs are mostly ____ and are ___-____enzymes but MMP-_, MMP-_ and MMP-__ are also found ____ and may act on intracellular proteins

They are synthesized as secreted or ___ ____ and processed to an active form by the ___ of an __-_____propeptide.

Interaction of a ____ residue in this peptide with the ___ moiety in the enzyme ___ ___ keeps the enzyme____.

Disruption of this interaction triggers ____.

There are several distinct subgroups based on preferential ____ or similar ____ domains:

____ that are active against ____ ____

____ that have high activity against ___ ____

____ that degrade ___ ____ ____ of the ECM

___ __ ____ (MT-MMPs) that are ____ molecules

The matrix metalloproteinases (MMPs) are a family of zinc metalloproteinases - these are Ca2+- and Zn2+-dependent endopeptidases that are active at neutral pH MMPs are mostly extracellular and are ECM-degrading enzymes but MMP-1, MMP-2 and MMP-11 are also found intracellularly and may act on intracellular proteins

They are synthesized as secreted or transmembrane proenzymes and processed to an active form by the removal of an amino-terminal propeptide. Interaction of a cysteine residue in this peptide with the zinc moiety in the enzyme active site keeps the enzyme inactive. Disruption of this interaction triggers activation There are several distinct subgroups based on preferential substrates or similar structural domains: Collagenases that are active against fibrillar collagen Gelatinases that have high activity against denatured collagens Stromelysins that degrade noncollagen components of the ECM Membrane-type MMPs (MT-MMPs) that are transmembrane molecules

2

Discovery of MMPs

The founding member of MMP family, collagenase, was identified in 1962 in tadpoles - found that tadpole tails during metamorphosis contained an enzyme that could degrade fibrillar collagen

Subsequently, an interstitial collagenase, collagenase-1 or MMP1, was found in diseased skin and synovium.

Further research led to the discovery of a family of 23 structurally related proteinases now referred to as the MMP family

The founding member of MMP family, collagenase, was identified in 1962 in tadpoles - found that tadpole tails during metamorphosis contained an enzyme that could degrade fibrillar collagen Subsequently, an interstitial collagenase, collagenase-1 or MMP1, was found in diseased skin and synovium. Further research led to the discovery of a family of 23 structurally related proteinases now referred to as the MMP family

3

Structure of MMPs

Mammalian MMPs share a conserved domain structure that consists of a ____ domain and an _____ __-domain (pro-peptide)

The pro-domain contains a conserved ____ residue that coordinates the active-site ___to inhibit catalysis

When the pro-domain is destabilized or removed, the active site becomes ___ to ___substrates

Most MMP-family members also contain a _____ domain, attached at their _ termini by a ___ ____.

The hemopexin domain encodes a ___ ____ ____ _____ structure that mediates:

  • ___ ____ interactions
  • substrate _____
  • ____ of the enzyme
  • protease ___ , ____ and _____

Structure of MMPs Mammalian MMPs share a conserved domain structure that consists of a catalytic domain and an autoinhibitory pro-domain (pro-peptide) The pro-domain contains a conserved Cys residue that coordinates the active-site zinc to inhibit catalysis When the pro-domain is destabilized or removed, the active site becomes available to cleave substrates Most MMP-family members also contain a hemopexin domain, attached at their C termini by a flexible hinge. The hemopexin domain encodes a four-bladed beta-propeller structure that mediates: protein–protein interactions substrate recognition activation of the enzyme protease localization, internalization and degradation

4

Classification of MMPs based on domain structure and substrate

1. _____and ____– consist of the basic structure of MMPs

2. Gelatinases - MMP2 and MMP9, have___ ____ type II ____ in their ____domains

3. Membrane type MMPs (MT-MMPs) - linked to the ___ ___ either by a _____ domain or by a ___________(GPI) linkage, attached to the _____domain

4. _____ - Minimal MMPs that lack the ___ and ___ domains

5. Others

1. Collagenases and stromelysins– consist of the basic structure of MMPs 2. Gelatinases - MMP2 and MMP9, have three fibronectin type II repeats in their catalytic domains 3. Membrane type MMPs (MT-MMPs) - linked to the plasma membrane either by a transmembrane domain or by a glycosylphosphatidylinositol (GPI) linkage, attached to the hemopexin domain 4. Matrilysins - Minimal MMPs that lack the hinge and hemopexin domains 5. Others

5

Cysteine-switch mechanism of MMP activation

Activation of MMPs is tightly regulated MMPs are synthesized as _____ (zymogens), which is an important regulatory step of MMP activity

Activation of pro-MMPs requires the disruption of the interaction between a ____ residue in their N-terminal pro-domain and the ___ ion at their catalytic center .

This can be achieved by either ___ modifying the ___ residue (for example by ___ ___ ___) or by proteolytically ____ a bait region in their pro-domain (black rectangle) by other proteases (even bacterial proteases).

Complete removal of the propeptide is often conducted in ___ by the action of the ___ ___ or by other ___ ___ thereby rendering the MMP fully active.

Activation of MMPs is tightly regulated MMPs are synthesized as proenzymes (zymogens), which is an important regulatory step of MMP activity Activation of pro-MMPs requires the disruption of the interaction between a cysteine residue in their N-terminal pro-domain and the zinc ion at their catalytic center . This can be achieved by either chemically modifying the cysteine residue (for example by reactive oxygen species) or by proteolytically cleaving a bait region in their pro-domain (black rectangle) by other proteases (even bacterial proteases). Complete removal of the propeptide is often conducted in trans by the action of the MMP intermediate or by other active MMPs thereby rendering the MMP fully active.

6

Regulation of MMP activity

MMPs can be regulated at the level of

  • RNA ____
  •  Protein____
  • ___, intracellular ____
  • Subcellular or extracellular ____,
  • Activation of the ____n form
  • Expression of their endogenous protein ____
  • Protease ____

MMPs can be regulated at the level of RNA transcription Protein synthesis Secretion, intracellular trafficking Subcellular or extracellular localization, Activation of the zymogen form Expression of their endogenous protein inhibitors Protease degradation

7

Endogenous inhibitors of MMPs

MMP activities are regulated by two major types of endogenous inhibitors: __-____ and ____ ____ ___ _____(TIMPs)

Human α2-macroglobin is a plasma glycoprotein that inhibits most proteinases by ____ the proteinase within the ____ and the complex is rapidly cleared by ___ ___ ___

TIMPs inhibit MMP activity by forming ___ affinity,____covalent, ______ _:_ complexes with the active forms of the proteinases

____ other proteins have also been reported to inhibit selected members of MMPs

MMP activities are regulated by two major types of endogenous inhibitors: α2-macroglobulin and tissue inhibitors of metalloproteinases (TIMPs) Human α2-macroglobin is a plasma glycoprotein that inhibits most proteinases by entrapping the proteinase within the macroglobulin and the complex is rapidly cleared by receptor-mediated endocytosis TIMPs inhibit MMP activity by forming high affinity, non-covalent, reversible 1:1 complexes with the active forms of the proteinases Several other proteins have also been reported to inhibit selected members of MMPs

8

Functions of MMPs

MMPs can cleave components of the extracellular matrix (ECM), resulting in increased ___ for cell or tissue___.

MMP proteolysis can generate specific ___ ___ that then___ in an ___ or ___ manner (Ex:Collagen II: fragment is a bone morphogenetic protein (BMP) antagonist)

MMPs can directly regulate ____ tissue ____through cleavage of ____ ____ or the ____ membrane

MMPs can ___ or ____ the action of latent ____molecules (Ex: growth factors), resulting in diverse cellular consequences

MMPs can ___ or___ the action of ___signaling molecules, resulting in changes in ___, cell __, ____ or cell ___

MMPs can cleave components of the extracellular matrix (ECM), resulting in increased space for cell or tissue movement MMP proteolysis can generate specific cleavage products that then signal in an autocrine or paracrine manner (Ex:Collagen II: fragment is a bone morphogenetic protein (BMP) antagonist) MMPs can directly regulate epithelial tissue architecture through cleavage of intercellular junctions or the basement membrane MMPs can activate or modify the action of latent signaling molecules (Ex: growth factors), resulting in diverse cellular consequences MMPs can deactivate or modify the action of active signaling molecules, resulting in changes in proliferation, cell death, differentiation or cell motility

9

Functions of MMPs in cancer progression

_____ MMP expression has been found in a number of different malignancies and expression has been related to tumor behavior and patient prognosis

Elevated MMP expression has been found in a number of different malignancies and expression has been related to tumor behavior and patient prognosis

10

Involvement of MMPs in oral diseases

MMPs and oral cancer

 

Several MMPs are ____ in oral tumors, in the ____ cells themselves as well as the ____ cells

Over expression of MMP_ and _ have been shown to induce tumor growth

Oral carcinoma patients with elevated MMP-_ and -_ activity have poor ____ than patients with low ____ tumor activities

Several MMPs are upregulated in oral tumors, in the tumor cells themselves as well as the surrounding cells Over expression of MMP2 and 9 have been shown to induce tumor growth Oral carcinoma patients with elevated MMP-2 and -9 activity have poor prognosis than patients with low gelatinase tumor activities

11

MMP___ and enamel formation

The ___, protein-rich extracellular matrix secreted by epithelial ____during the secretory stage develops into hard enamel tissue

Before tooth eruption, digestion of enamel proteins by _____ and ____ by ___ provide ____ for ___ expansion of the tooth hydroxyapatite crystalline and facilitates tooth ___.

 KLK4 is secreted by ____ as a ___

MMP20, also known as _____, cleaves __-KLK4 to active KLK4

Reduction or absence of KLK4 activity due to mutations in MMP20 has been shown to lead to ____-related pathologies, such as ___ ___

Note - MMPs play an important role in the matrix remodeling that takes place during _____ as well.

The soft, protein-rich extracellular matrix secreted by epithelial ameloblasts during the secretory stage develops into hard enamel tissue Before tooth eruption, digestion of enamel proteins by kallikrein 4 (KLK4) and reabsorption by ameloblasts provide space for lateral expansion of the tooth hydroxyapatite crystalline and facilitates tooth hardening KLK4 is secreted by ameloblasts as a zymogen MMP20, also known as enamelysin, cleaves Pro-KLK4 to active KLK4 Reduction or absence of KLK4 activity due to mutations in MMP20 has been shown to lead to amelogenesis-related pathologies, such as amelogenesis imperfecta Note - MMPs play an important role in the matrix remodeling that takes place during dentinogenesis as well.

12

MMPs and caries

Because enamel is highly____, the enamel carious process involves ____ reactions, where the mineral is dissolved by ____ liberated by cariogenic bacteria

In contrast, the dentin is ___ mineralized and contains an abundant ____ material (20%), mainly composed of type __ collagen.

Therefore, the dentin carious process strongly differs from the enamel carious process

MMPs have been suggested to play an important role in the ____ of dentin organic matrix following ____ by bacterial___ and, therefore, in the control or progression of carious decay

Host-derived MMPs can originate both from ___ and from ___. They may be activated by an ___ pH brought about by the bacteria or by bacterial ____

Once the local pH is ____ by ____ buffer systems, activated MMPs degrade the ____ dentin matrix

MMPs and caries Because enamel is highly mineralized, the enamel carious process involves physicochemical reactions, where the mineral is dissolved by acids liberated by cariogenic bacteria In contrast, the dentin is less mineralized and contains an abundant organic material (20%), mainly composed of type I collagen. Therefore, the dentin carious process strongly differs from the enamel carious process MMPs have been suggested to play an important role in the destruction of dentin organic matrix following demineralization by bacterial acids and, therefore, in the control or progression of carious decay Host-derived MMPs can originate both from saliva and from dentin. They may be activated by an acidic pH brought about by the bacteria or by bacterial proteases Once the local pH is neutralized by salivary buffer systems, activated MMPs degrade the demineralized dentin matrix

13

MMPs and periodontitis

___ and____degradation are important processes in the pathogenesis of periodontitis

Increasing evidence indicates that host MMPs, mainly MMP_ and MMP_, are essential in these processes

There is evidence that the level of MMP-_ in the _____ of periodontal pockets correlates with clinical periodontal parameters

Chairside tests for qualitative detection of MMP8 in crevicular fluid has been used for early ____ of periodontitis

ECM and dentin degradation are important processes in the pathogenesis of periodontitis Increasing evidence indicates that host MMPs, mainly MMP8 and MMP9, are essential in these processes There is evidence that the level of MMP-8 in the gingival crevicular fluid (GCF) of periodontal pockets correlates with clinical periodontal parameters Chairside tests for qualitative detection of MMP8 in crevicular fluid has been used for early detection of periodontitis