Journal: Molecular Vision

Article Title: Matrix metalloproteinases in recurrent corneal melting associated with primary Sjörgen’s syndrome

doi:

Figure Lengend Snippet: Matrix metalloproteinase detecting antibodies used for indirect immunohistochemistry.

Article Snippet: Monoclonal mouse anti-human MMP 2 , MAB13431 , 1:350 , Chemicon Intl. Inc..

Techniques: Immunohistochemistry, Concentration Assay

Journal: Molecular Vision

Article Title: Matrix metalloproteinases in recurrent corneal melting associated with primary Sjörgen’s syndrome

doi:

Figure Lengend Snippet: The immunohistochemical localization of individual MMPs in the corneal specimens obtained from two patients with pSS (S1-8 and S9-11 respectively) and the average values of immunohistochemical staining in all melted specimens (S) and controls (C).

Article Snippet: Monoclonal mouse anti-human MMP 2 , MAB13431 , 1:350 , Chemicon Intl. Inc..

Techniques: Immunohistochemical staining, Staining

Journal: Molecular Vision

Article Title: Matrix metalloproteinases in recurrent corneal melting associated with primary Sjörgen’s syndrome

doi:

Figure Lengend Snippet: Gelatin zymography of matrix metalloproteinases 2 and 9 in melted and control corneas. Melted specimens (A) S1 and S2 showed extremely high levels, and specimens S7, S9, and S11 considerable levels, of both the proenzyme (67 KDa band) and the active form of MMP 2 (62 kDa band). Levels of MMP 9 proenzyme (95 kDa band) and the active form (82 kDa band) were extremely high in S4 and S6-S11, and prominent in S5. Weak bands for both MMP 9 forms were found in specimens S1-S3. In control samples (B) , a moderate level of the MMP 2 proenzyme was present in all specimens, whereas the active form of MMP 2 was either not present or very faint, except in samples C1, C3, and C11. As for MMP 9 proenzyme, only C3 and C7 showed faint bands.

Article Snippet: Monoclonal mouse anti-human MMP 2 , MAB13431 , 1:350 , Chemicon Intl. Inc..

Techniques: Zymography

Journal: Cancers

Article Title: Ciliary Neurotrophic Factor Modulates Multiple Downstream Signaling Pathways in Prostate Cancer Inhibiting Cell Invasiveness

doi: 10.3390/cancers14235917

Figure Lengend Snippet: Antibodies used in this study.

Article Snippet: mAb Mouse anti-human MMP-2 (#436000) , // , 1:500 , // , Thermo Fisher Scientific, Waltham, MA, USA.

Techniques:

Journal: PLoS ONE

Article Title: Phosphorylation Status of 72 kDa MMP-2 Determines Its Structure and Activity in Response to Peroxynitrite

doi: 10.1371/journal.pone.0071794

Figure Lengend Snippet: (A) Domain structure, NetPhos predicted phosphorylation sites, cysteine residues and disulphide bonds of human MMP-2. Vertical white, grey and black lines that extend above the demarcated horizontal threshold indicate putative amino acid phosphorylation sites, whether serine, threonine, or tyrosine, respectively. Red vertical lines indicate all cysteine residues present in the MMP-2 sequence, and blue horizontal lines represent the disulphide bonds of human MMP-2. (B) Crystal structure of MMP-2 (PDB ID: 1CK7) is shown in cartoon representation. The pro-peptide (hot pink), collagenase-like domain 1 (deep teal), collagenase-binding domain (forest green), collagenase-like domain 2 (pale green) and hemopexin-like domain (yellow) are shown in the indicated colours. Zn 2+ ions are indicated as purple spheres. Potential phosphorylation sites (serine, threonine and tyrosine) are shown in Corey-Pauling-Koltun (CPK)/space filling representation, with carbon and oxygen atoms of the side chains in grey and red, respectively.

Article Snippet: The following were purchased from the sources indicated: alkaline phosphatase from bovine intestinal mucosa (Sigma-Aldrich, St. Louis MO, USA), protein-A Sepharose bead suspension (BioVision, Mountain View CA, USA), mouse monoclonal IgG 1 and mouse monoclonal glutathione antibodies (Abcam, Cambridge MA, USA), mouse monoclonal anti-human MMP-2 antibody (Millipore, Temecula CA, USA), secondary horseradish peroxidase-conjugated antibody (Transduction Laboratories, Mississauga, ON), ECL Plus (Amersham, Buckinghamshire, UK); OmniMMP® fluorogenic peptide substrate (Enzo Life Sciences, Plymouth Meeting PA, USA); Phos-tag™ acrylamide (NARD Institute Ltd., Amagasaki, Japan); Precision Plus protein dual color molecular weight standards and Coomassie Brilliant Blue R-250 (Bio-Rad, Hercules CA, USA).

Techniques: Sequencing, Binding Assay

Journal: PLoS ONE

Article Title: Phosphorylation Status of 72 kDa MMP-2 Determines Its Structure and Activity in Response to Peroxynitrite

doi: 10.1371/journal.pone.0071794

Figure Lengend Snippet: (A) In representative Coomassie blue stained SDS-PAGE gels, both native (phosphorylated, +P) and dephosphorylated (−P) 72 kDa MMP-2 run with the same pattern, showing bands only at 72 kDa (left). However, when run on the Phos-tag acrylamide gel, a more slowly migrating band also appeared, indicating that a portion of MMP-2 is phosphorylated. Phos-tag gel revealed at least two different phosphorylation states of 72 kDa MMP-2 and confirmed its dephosphorylation after treatment with alkaline phosphatase (right, N = 4). (B) The ratio of band intensity of total MMP-2 measured by SDS-PAGE (left), as well as ratio of higher to lower band intensities measured in the Phos-tag gel (right, mean ± SEM, N = 4), comparing phosphorylated to dephosphorylated samples, is demonstrated in the bar graphs below. (C) CD spectra of hrMMP-2. Far UV range CD spectrum of native (phosphorylated, black line) and dephosphorylated (red line).

Article Snippet: The following were purchased from the sources indicated: alkaline phosphatase from bovine intestinal mucosa (Sigma-Aldrich, St. Louis MO, USA), protein-A Sepharose bead suspension (BioVision, Mountain View CA, USA), mouse monoclonal IgG 1 and mouse monoclonal glutathione antibodies (Abcam, Cambridge MA, USA), mouse monoclonal anti-human MMP-2 antibody (Millipore, Temecula CA, USA), secondary horseradish peroxidase-conjugated antibody (Transduction Laboratories, Mississauga, ON), ECL Plus (Amersham, Buckinghamshire, UK); OmniMMP® fluorogenic peptide substrate (Enzo Life Sciences, Plymouth Meeting PA, USA); Phos-tag™ acrylamide (NARD Institute Ltd., Amagasaki, Japan); Precision Plus protein dual color molecular weight standards and Coomassie Brilliant Blue R-250 (Bio-Rad, Hercules CA, USA).

Techniques: Staining, SDS Page, Acrylamide Gel Assay, De-Phosphorylation Assay

Journal: PLoS ONE

Article Title: Phosphorylation Status of 72 kDa MMP-2 Determines Its Structure and Activity in Response to Peroxynitrite

doi: 10.1371/journal.pone.0071794

Figure Lengend Snippet: S-glutathiolation of phosphorylated (A) or dephosphorylated (B) MMP-2, treated with 0.3 µM ONOO − in the presence or absence of 30 µM GSH was determined by immunoprecipitation with glutathione (GSH) antibody, or IgG as a control, followed by Western blot for MMP-2. M, MMP-2; O, 0.3 µM ONOO − ; G, 30 µM GSH; D, 1 mM dithiothreitol.

Article Snippet: The following were purchased from the sources indicated: alkaline phosphatase from bovine intestinal mucosa (Sigma-Aldrich, St. Louis MO, USA), protein-A Sepharose bead suspension (BioVision, Mountain View CA, USA), mouse monoclonal IgG 1 and mouse monoclonal glutathione antibodies (Abcam, Cambridge MA, USA), mouse monoclonal anti-human MMP-2 antibody (Millipore, Temecula CA, USA), secondary horseradish peroxidase-conjugated antibody (Transduction Laboratories, Mississauga, ON), ECL Plus (Amersham, Buckinghamshire, UK); OmniMMP® fluorogenic peptide substrate (Enzo Life Sciences, Plymouth Meeting PA, USA); Phos-tag™ acrylamide (NARD Institute Ltd., Amagasaki, Japan); Precision Plus protein dual color molecular weight standards and Coomassie Brilliant Blue R-250 (Bio-Rad, Hercules CA, USA).

Techniques: Immunoprecipitation, Western Blot

Journal: PLoS ONE

Article Title: Phosphorylation Status of 72 kDa MMP-2 Determines Its Structure and Activity in Response to Peroxynitrite

doi: 10.1371/journal.pone.0071794

Figure Lengend Snippet: Mass spectrometry data obtained for native and dephosphorylated MMP-2 treated with 0.3 µM ONOO − and 30 µM GSH showing modifications in Cys residues.

Article Snippet: The following were purchased from the sources indicated: alkaline phosphatase from bovine intestinal mucosa (Sigma-Aldrich, St. Louis MO, USA), protein-A Sepharose bead suspension (BioVision, Mountain View CA, USA), mouse monoclonal IgG 1 and mouse monoclonal glutathione antibodies (Abcam, Cambridge MA, USA), mouse monoclonal anti-human MMP-2 antibody (Millipore, Temecula CA, USA), secondary horseradish peroxidase-conjugated antibody (Transduction Laboratories, Mississauga, ON), ECL Plus (Amersham, Buckinghamshire, UK); OmniMMP® fluorogenic peptide substrate (Enzo Life Sciences, Plymouth Meeting PA, USA); Phos-tag™ acrylamide (NARD Institute Ltd., Amagasaki, Japan); Precision Plus protein dual color molecular weight standards and Coomassie Brilliant Blue R-250 (Bio-Rad, Hercules CA, USA).

Techniques: Mass Spectrometry

Journal: PLoS ONE

Article Title: Phosphorylation Status of 72 kDa MMP-2 Determines Its Structure and Activity in Response to Peroxynitrite

doi: 10.1371/journal.pone.0071794

Figure Lengend Snippet: Proteolysis of fluorogenic Omni MMP substrate by ONOO − -treated, native (+P, left panel) or dephosphorylated (−P, right panel) 72 kDa MMP-2, in the presence of 30 µM GSH. Mean ± SEM, N = 7–8/group. *p<0.05 versus 0 µM ONOO − . DPN, decomposed peroxynitrite.

Article Snippet: The following were purchased from the sources indicated: alkaline phosphatase from bovine intestinal mucosa (Sigma-Aldrich, St. Louis MO, USA), protein-A Sepharose bead suspension (BioVision, Mountain View CA, USA), mouse monoclonal IgG 1 and mouse monoclonal glutathione antibodies (Abcam, Cambridge MA, USA), mouse monoclonal anti-human MMP-2 antibody (Millipore, Temecula CA, USA), secondary horseradish peroxidase-conjugated antibody (Transduction Laboratories, Mississauga, ON), ECL Plus (Amersham, Buckinghamshire, UK); OmniMMP® fluorogenic peptide substrate (Enzo Life Sciences, Plymouth Meeting PA, USA); Phos-tag™ acrylamide (NARD Institute Ltd., Amagasaki, Japan); Precision Plus protein dual color molecular weight standards and Coomassie Brilliant Blue R-250 (Bio-Rad, Hercules CA, USA).

Techniques:

Journal: PLoS ONE

Article Title: Phosphorylation Status of 72 kDa MMP-2 Determines Its Structure and Activity in Response to Peroxynitrite

doi: 10.1371/journal.pone.0071794

Figure Lengend Snippet: (A) Representative time-dependent troponin I (TnI) hydrolysis by 0.3 µM ONOO − - treated, native (upper panel) or dephosphorylated (lower panel) 72 kDa MMP-2, in the presence of 30 µM GSH, following different incubation times (30, 60, or 120 min) at 37°C with TnI. Representative Coomassie blue stained SDS-PAGE gels. (B) Quantitative analysis of TnI hydrolysis by native (left) or dephosphorylated (right) 72 kDa MMP-2, treated with different concentrations of ONOO − (0–0.3 µM) or DPN, in the presence of 30 µM GSH. Incubation for 120 min at 37°C. Mean ± SEM, N = 4–7/group. * p<0.05 compared with control (C, TnI alone). DPN, decomposed peroxynitrite.

Article Snippet: The following were purchased from the sources indicated: alkaline phosphatase from bovine intestinal mucosa (Sigma-Aldrich, St. Louis MO, USA), protein-A Sepharose bead suspension (BioVision, Mountain View CA, USA), mouse monoclonal IgG 1 and mouse monoclonal glutathione antibodies (Abcam, Cambridge MA, USA), mouse monoclonal anti-human MMP-2 antibody (Millipore, Temecula CA, USA), secondary horseradish peroxidase-conjugated antibody (Transduction Laboratories, Mississauga, ON), ECL Plus (Amersham, Buckinghamshire, UK); OmniMMP® fluorogenic peptide substrate (Enzo Life Sciences, Plymouth Meeting PA, USA); Phos-tag™ acrylamide (NARD Institute Ltd., Amagasaki, Japan); Precision Plus protein dual color molecular weight standards and Coomassie Brilliant Blue R-250 (Bio-Rad, Hercules CA, USA).

Techniques: Incubation, Staining, SDS Page