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Rac1 promotes Rab5-dependent endocytosis of Crb. (A-E) Embryos were co-stained for Crb (red in the merged images) and Arm (green). Panels depict a surface view of the ventral ectoderm of the following embryos: Control (A: da-GAL4 embryo incubated in DMSO), Rac1 V12 (B: da-GAL4/UAS-Rac1 V12 embryo incubated in DMSO), Rac1 V12 <t>Leupeptin</t> (C: da-GAL4/UAS-Rac1 V12 embryo treated with the lysosomal proteolysis inhibitor leupeptin), Rac1 V12 Dynasore (D: da-GAL4/UAS-Rac1 V12 embryo treated with the dynamin inhibitor dynasore), Rac1 V12 Rab5 S43N ( da-GAL4/UAS-Rac1 V12 , UAS-Rab5 S43N ). Images are representative results of three independent experiments. Scale bar: 10 µm. C, arrows point to intracellular puncta; D,E, arrows indicate cell-cell borders. (F-H) Stage 16 Rab5 S43N -expressing embryos stained for 2A12 (F), Serp (G) or Verm (H). (I) 2A12 staining performed on an embryo expressing Rab5 S43N in the trachea and heterozygous for the null allele crb 11A22 ( btl-GAL4; UAS-Rab5 S43N / crb 11A22 ). Scale bar: 100 µm. (J) Histogram showing the quantification of the dorsal trunk length in Rab5 S43N ( btl-GAL4; UAS-Rab5 S43N ) and Rab5 S43N , crb /+ ( btl-GAL4; UAS-Rab5 S43N , crb 11A22 /+ ) embryos relative to dorsal trunk length of control animals ( btl-GAL4 ). n =15 embryos for each genotype, taken from at least three independent experiments. Bars represent mean±s.d. A two-tailed t -test was used to evaluate the statistical significance; ** P
Leupeptin, supplied by BioShop, used in various techniques. Bioz Stars score: 93/100, based on 14 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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1) Product Images from "Rac1 controls epithelial tube length through the apical secretion and polarity pathways"

Article Title: Rac1 controls epithelial tube length through the apical secretion and polarity pathways

Journal: Biology Open

doi: 10.1242/bio.015727

Rac1 promotes Rab5-dependent endocytosis of Crb. (A-E) Embryos were co-stained for Crb (red in the merged images) and Arm (green). Panels depict a surface view of the ventral ectoderm of the following embryos: Control (A: da-GAL4 embryo incubated in DMSO), Rac1 V12 (B: da-GAL4/UAS-Rac1 V12 embryo incubated in DMSO), Rac1 V12 Leupeptin (C: da-GAL4/UAS-Rac1 V12 embryo treated with the lysosomal proteolysis inhibitor leupeptin), Rac1 V12 Dynasore (D: da-GAL4/UAS-Rac1 V12 embryo treated with the dynamin inhibitor dynasore), Rac1 V12 Rab5 S43N ( da-GAL4/UAS-Rac1 V12 , UAS-Rab5 S43N ). Images are representative results of three independent experiments. Scale bar: 10 µm. C, arrows point to intracellular puncta; D,E, arrows indicate cell-cell borders. (F-H) Stage 16 Rab5 S43N -expressing embryos stained for 2A12 (F), Serp (G) or Verm (H). (I) 2A12 staining performed on an embryo expressing Rab5 S43N in the trachea and heterozygous for the null allele crb 11A22 ( btl-GAL4; UAS-Rab5 S43N / crb 11A22 ). Scale bar: 100 µm. (J) Histogram showing the quantification of the dorsal trunk length in Rab5 S43N ( btl-GAL4; UAS-Rab5 S43N ) and Rab5 S43N , crb /+ ( btl-GAL4; UAS-Rab5 S43N , crb 11A22 /+ ) embryos relative to dorsal trunk length of control animals ( btl-GAL4 ). n =15 embryos for each genotype, taken from at least three independent experiments. Bars represent mean±s.d. A two-tailed t -test was used to evaluate the statistical significance; ** P
Figure Legend Snippet: Rac1 promotes Rab5-dependent endocytosis of Crb. (A-E) Embryos were co-stained for Crb (red in the merged images) and Arm (green). Panels depict a surface view of the ventral ectoderm of the following embryos: Control (A: da-GAL4 embryo incubated in DMSO), Rac1 V12 (B: da-GAL4/UAS-Rac1 V12 embryo incubated in DMSO), Rac1 V12 Leupeptin (C: da-GAL4/UAS-Rac1 V12 embryo treated with the lysosomal proteolysis inhibitor leupeptin), Rac1 V12 Dynasore (D: da-GAL4/UAS-Rac1 V12 embryo treated with the dynamin inhibitor dynasore), Rac1 V12 Rab5 S43N ( da-GAL4/UAS-Rac1 V12 , UAS-Rab5 S43N ). Images are representative results of three independent experiments. Scale bar: 10 µm. C, arrows point to intracellular puncta; D,E, arrows indicate cell-cell borders. (F-H) Stage 16 Rab5 S43N -expressing embryos stained for 2A12 (F), Serp (G) or Verm (H). (I) 2A12 staining performed on an embryo expressing Rab5 S43N in the trachea and heterozygous for the null allele crb 11A22 ( btl-GAL4; UAS-Rab5 S43N / crb 11A22 ). Scale bar: 100 µm. (J) Histogram showing the quantification of the dorsal trunk length in Rab5 S43N ( btl-GAL4; UAS-Rab5 S43N ) and Rab5 S43N , crb /+ ( btl-GAL4; UAS-Rab5 S43N , crb 11A22 /+ ) embryos relative to dorsal trunk length of control animals ( btl-GAL4 ). n =15 embryos for each genotype, taken from at least three independent experiments. Bars represent mean±s.d. A two-tailed t -test was used to evaluate the statistical significance; ** P

Techniques Used: Staining, Incubation, Expressing, Two Tailed Test

2) Product Images from "ROCK inhibitors upregulate the neuroprotective Parkin-mediated mitophagy pathway"

Article Title: ROCK inhibitors upregulate the neuroprotective Parkin-mediated mitophagy pathway

Journal: Nature Communications

doi: 10.1038/s41467-019-13781-3

Targeting of mitochondria to lysosomes is increased by SR3677. a HeLa cells were co-transfected with Cerulean-Parkin and RG-OMP25. After 24 h, cells were pre-treated with either 0.5 µM SR3677 or DMSO for 2 h prior to induction of mitophagy with 10 µM CCCP treatment, in combination with E-64 and leupeptin. Red-only signal represents mitochondria localized to lysosomes, where GFP signal is quenched. Scale bars, 10 µm. b Quantification of the percentage of red-only mitochondrial area divided by the total non-background area ( n = 4 independent experiments). P -values were determined by one-tailed paired Student’s t -test. c Seven-day-old TH-GAL4 > UAS-mitoQC male flies were placed into vials containing the indicated treatments. Representative images of the dopaminergic neurons of TH-GAL4 > UAS-mitoQC flies following feeding on fly food supplemented with H 2 O, 0.5 mM SR3677 (SR) or H 2 O/SR3677 combined with 5 mM paraquat (PQ). Scale bars, 10 µm. d Quantification of the percentage of red-only mitochondrial area divided by the total non-background area, averaged across 0.8-µm z-stacks. Data are expressed as mean ± s.e.m ( n = 4 independent experiments). P -values were determined by one-tailed paired Student’s t -test, * P
Figure Legend Snippet: Targeting of mitochondria to lysosomes is increased by SR3677. a HeLa cells were co-transfected with Cerulean-Parkin and RG-OMP25. After 24 h, cells were pre-treated with either 0.5 µM SR3677 or DMSO for 2 h prior to induction of mitophagy with 10 µM CCCP treatment, in combination with E-64 and leupeptin. Red-only signal represents mitochondria localized to lysosomes, where GFP signal is quenched. Scale bars, 10 µm. b Quantification of the percentage of red-only mitochondrial area divided by the total non-background area ( n = 4 independent experiments). P -values were determined by one-tailed paired Student’s t -test. c Seven-day-old TH-GAL4 > UAS-mitoQC male flies were placed into vials containing the indicated treatments. Representative images of the dopaminergic neurons of TH-GAL4 > UAS-mitoQC flies following feeding on fly food supplemented with H 2 O, 0.5 mM SR3677 (SR) or H 2 O/SR3677 combined with 5 mM paraquat (PQ). Scale bars, 10 µm. d Quantification of the percentage of red-only mitochondrial area divided by the total non-background area, averaged across 0.8-µm z-stacks. Data are expressed as mean ± s.e.m ( n = 4 independent experiments). P -values were determined by one-tailed paired Student’s t -test, * P

Techniques Used: Transfection, One-tailed Test

3) Product Images from "Inhibition of Outer Membrane Proteases of the Omptin Family by Aprotinin"

Article Title: Inhibition of Outer Membrane Proteases of the Omptin Family by Aprotinin

Journal: Infection and Immunity

doi: 10.1128/IAI.00136-15

Inhibition of CroP activity by aprotinin. (A) Aprotinin inhibits cleavage of the FRET substrate by purified CroP. FRET assays were performed with 10 nM CroP in PBS (black) or in the presence of EDTA (2.5 mM, green), PMSF (2.5 mM, purple), leupeptin (10
Figure Legend Snippet: Inhibition of CroP activity by aprotinin. (A) Aprotinin inhibits cleavage of the FRET substrate by purified CroP. FRET assays were performed with 10 nM CroP in PBS (black) or in the presence of EDTA (2.5 mM, green), PMSF (2.5 mM, purple), leupeptin (10

Techniques Used: Inhibition, Activity Assay, Purification

Related Articles

Activation Assay:

Article Title: Hsc70 chaperone activity underlies Trio GEF function in axon growth and guidance induced by netrin-1
Article Snippet: .. Rac1 activation assay Transfected HEK293 cells were serum starved overnight and then lysed in buffer containing 25 mM Hepes, pH 7.5, 1% NP-40, 10 mM MgCl2 , 100 mM NaCl, 5% glycerol, 1 mM PMSF, and 1 µg/ml aprotinin and leupeptin (BioShop). ..

Incubation:

Article Title: Rac1 controls epithelial tube length through the apical secretion and polarity pathways
Article Snippet: .. Pharmacological treatment of embryos Dechorionated embryos were incubated in the dark at 25°C in a solution of 0.9% NaCl (under an octane phase) supplemented with DMSO (control), 0.8 mM Dynasore (30 min; Cedarlane) or 0.1 mg/ml leupeptin (3 h; Bioshop). ..

Article Title: The mammalian CTLH complex is an E3 ubiquitin ligase that targets its subunit muskelin for degradation
Article Snippet: .. Cells were lysed in denaturing buffer (50 mM Tris, pH 7.5, 150 mM NaCl, 1% Triton, 1% SDS, 1 mM Na3VO4, 10 mM NaF, 1 mM phenylmethylsulfonyl fluoride (PMSF), 1 μg/ml of aprotinin, 10 μg/ml of peptatin, 1 μg/ml of leupeptin and 25 mM NEM (N-Ethylamalide, Bioshop Canada, Burlington, ON, Canada)), passed through a 23 G needle ten times and incubated on ice for 30 minutes. .. For immunoprecipitation, lysates were diluted 1:10 in buffer A (50 mM Tris, pH 7.5, 150 mM NaCl, 1% Triton, 1 mM Na3VO4, 10 mM NaF and 25 mM NEM) and incubated with anti-FLAG (M2; F1804; Sigma-Aldrich) for 2 hours at 4 °C, followed by incubation with Dynabeads Protein G for 1 hour.

other:

Article Title: Differential Regulation of Endosomal GPCR/β-Arrestin Complexes and Trafficking by MAPK *
Article Snippet: Phenylmethyl sulfonyl fluoride (PMSF), aprotinin, leupeptin, and pepstatin were from Bioshop (Burlington, Canada).

Article Title: ROCK inhibitors upregulate the neuroprotective Parkin-mediated mitophagy pathway
Article Snippet: Chemicals The following chemicals were used in our study: CCCP (Sigma, C2759), SR3677 (Tocris, 3667/10), Y27632 (Millipore, 688000), Y39983 (MedChemExpress, HY-10069), paraquat (Sigma, 36541), E-64 (BioShop, EEL640.1), leupeptin (BioShop, Leu001.50), geneticin (Gibco, 11811023), puromycin (BioShop, PUR333.10), retinoic acid (Calbiochem, 554720), brain-derived neurotrophic factor (Gibco, PHC7074) and chloroquine (Bioshop, CHL919).

Transfection:

Article Title: Hsc70 chaperone activity underlies Trio GEF function in axon growth and guidance induced by netrin-1
Article Snippet: .. Rac1 activation assay Transfected HEK293 cells were serum starved overnight and then lysed in buffer containing 25 mM Hepes, pH 7.5, 1% NP-40, 10 mM MgCl2 , 100 mM NaCl, 5% glycerol, 1 mM PMSF, and 1 µg/ml aprotinin and leupeptin (BioShop). ..

Lysis:

Article Title: Identification of Differentially Regulated Secretome Components During Skeletal Myogenesis *
Article Snippet: .. The supernatant was discarded and the remaining pellet was resuspended in 200 μl lysis buffer composed of 50 m m Tris-HCl (Bioshop), 150 m m NaCl (Bioshop), 0.5% Nonidet P-40 (BioRad), 2 m m EDTA (Bioshop), 100 m m NaF (Sigma), 10 m m Na2 HPO4 (Bioshop), 1 m m Na3 VO4 (Sigma), 1 m m PMSF (Bioshop), 1 μg/ml leupeptin (Bioshop), 1 μg/ml aprotinin (Bioshop), and 1 μg/ml pepstatin A (Bioshop). .. To examine the expression pattern of the secretome during myogenesis, CM were collected from light-labeled MBs and heavy-labeled MTs cultured in serum-free DM for 24 h and 120 h, respectively (see Step 2 of ).

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    BioShop leupeptin
    Rac1 promotes Rab5-dependent endocytosis of Crb. (A-E) Embryos were co-stained for Crb (red in the merged images) and Arm (green). Panels depict a surface view of the ventral ectoderm of the following embryos: Control (A: da-GAL4 embryo incubated in DMSO), Rac1 V12 (B: da-GAL4/UAS-Rac1 V12 embryo incubated in DMSO), Rac1 V12 <t>Leupeptin</t> (C: da-GAL4/UAS-Rac1 V12 embryo treated with the lysosomal proteolysis inhibitor leupeptin), Rac1 V12 Dynasore (D: da-GAL4/UAS-Rac1 V12 embryo treated with the dynamin inhibitor dynasore), Rac1 V12 Rab5 S43N ( da-GAL4/UAS-Rac1 V12 , UAS-Rab5 S43N ). Images are representative results of three independent experiments. Scale bar: 10 µm. C, arrows point to intracellular puncta; D,E, arrows indicate cell-cell borders. (F-H) Stage 16 Rab5 S43N -expressing embryos stained for 2A12 (F), Serp (G) or Verm (H). (I) 2A12 staining performed on an embryo expressing Rab5 S43N in the trachea and heterozygous for the null allele crb 11A22 ( btl-GAL4; UAS-Rab5 S43N / crb 11A22 ). Scale bar: 100 µm. (J) Histogram showing the quantification of the dorsal trunk length in Rab5 S43N ( btl-GAL4; UAS-Rab5 S43N ) and Rab5 S43N , crb /+ ( btl-GAL4; UAS-Rab5 S43N , crb 11A22 /+ ) embryos relative to dorsal trunk length of control animals ( btl-GAL4 ). n =15 embryos for each genotype, taken from at least three independent experiments. Bars represent mean±s.d. A two-tailed t -test was used to evaluate the statistical significance; ** P
    Leupeptin, supplied by BioShop, used in various techniques. Bioz Stars score: 93/100, based on 14 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/leupeptin/product/BioShop
    Average 93 stars, based on 14 article reviews
    Price from $9.99 to $1999.99
    leupeptin - by Bioz Stars, 2020-07
    93/100 stars
      Buy from Supplier

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    Rac1 promotes Rab5-dependent endocytosis of Crb. (A-E) Embryos were co-stained for Crb (red in the merged images) and Arm (green). Panels depict a surface view of the ventral ectoderm of the following embryos: Control (A: da-GAL4 embryo incubated in DMSO), Rac1 V12 (B: da-GAL4/UAS-Rac1 V12 embryo incubated in DMSO), Rac1 V12 Leupeptin (C: da-GAL4/UAS-Rac1 V12 embryo treated with the lysosomal proteolysis inhibitor leupeptin), Rac1 V12 Dynasore (D: da-GAL4/UAS-Rac1 V12 embryo treated with the dynamin inhibitor dynasore), Rac1 V12 Rab5 S43N ( da-GAL4/UAS-Rac1 V12 , UAS-Rab5 S43N ). Images are representative results of three independent experiments. Scale bar: 10 µm. C, arrows point to intracellular puncta; D,E, arrows indicate cell-cell borders. (F-H) Stage 16 Rab5 S43N -expressing embryos stained for 2A12 (F), Serp (G) or Verm (H). (I) 2A12 staining performed on an embryo expressing Rab5 S43N in the trachea and heterozygous for the null allele crb 11A22 ( btl-GAL4; UAS-Rab5 S43N / crb 11A22 ). Scale bar: 100 µm. (J) Histogram showing the quantification of the dorsal trunk length in Rab5 S43N ( btl-GAL4; UAS-Rab5 S43N ) and Rab5 S43N , crb /+ ( btl-GAL4; UAS-Rab5 S43N , crb 11A22 /+ ) embryos relative to dorsal trunk length of control animals ( btl-GAL4 ). n =15 embryos for each genotype, taken from at least three independent experiments. Bars represent mean±s.d. A two-tailed t -test was used to evaluate the statistical significance; ** P

    Journal: Biology Open

    Article Title: Rac1 controls epithelial tube length through the apical secretion and polarity pathways

    doi: 10.1242/bio.015727

    Figure Lengend Snippet: Rac1 promotes Rab5-dependent endocytosis of Crb. (A-E) Embryos were co-stained for Crb (red in the merged images) and Arm (green). Panels depict a surface view of the ventral ectoderm of the following embryos: Control (A: da-GAL4 embryo incubated in DMSO), Rac1 V12 (B: da-GAL4/UAS-Rac1 V12 embryo incubated in DMSO), Rac1 V12 Leupeptin (C: da-GAL4/UAS-Rac1 V12 embryo treated with the lysosomal proteolysis inhibitor leupeptin), Rac1 V12 Dynasore (D: da-GAL4/UAS-Rac1 V12 embryo treated with the dynamin inhibitor dynasore), Rac1 V12 Rab5 S43N ( da-GAL4/UAS-Rac1 V12 , UAS-Rab5 S43N ). Images are representative results of three independent experiments. Scale bar: 10 µm. C, arrows point to intracellular puncta; D,E, arrows indicate cell-cell borders. (F-H) Stage 16 Rab5 S43N -expressing embryos stained for 2A12 (F), Serp (G) or Verm (H). (I) 2A12 staining performed on an embryo expressing Rab5 S43N in the trachea and heterozygous for the null allele crb 11A22 ( btl-GAL4; UAS-Rab5 S43N / crb 11A22 ). Scale bar: 100 µm. (J) Histogram showing the quantification of the dorsal trunk length in Rab5 S43N ( btl-GAL4; UAS-Rab5 S43N ) and Rab5 S43N , crb /+ ( btl-GAL4; UAS-Rab5 S43N , crb 11A22 /+ ) embryos relative to dorsal trunk length of control animals ( btl-GAL4 ). n =15 embryos for each genotype, taken from at least three independent experiments. Bars represent mean±s.d. A two-tailed t -test was used to evaluate the statistical significance; ** P

    Article Snippet: Pharmacological treatment of embryos Dechorionated embryos were incubated in the dark at 25°C in a solution of 0.9% NaCl (under an octane phase) supplemented with DMSO (control), 0.8 mM Dynasore (30 min; Cedarlane) or 0.1 mg/ml leupeptin (3 h; Bioshop).

    Techniques: Staining, Incubation, Expressing, Two Tailed Test

    Targeting of mitochondria to lysosomes is increased by SR3677. a HeLa cells were co-transfected with Cerulean-Parkin and RG-OMP25. After 24 h, cells were pre-treated with either 0.5 µM SR3677 or DMSO for 2 h prior to induction of mitophagy with 10 µM CCCP treatment, in combination with E-64 and leupeptin. Red-only signal represents mitochondria localized to lysosomes, where GFP signal is quenched. Scale bars, 10 µm. b Quantification of the percentage of red-only mitochondrial area divided by the total non-background area ( n = 4 independent experiments). P -values were determined by one-tailed paired Student’s t -test. c Seven-day-old TH-GAL4 > UAS-mitoQC male flies were placed into vials containing the indicated treatments. Representative images of the dopaminergic neurons of TH-GAL4 > UAS-mitoQC flies following feeding on fly food supplemented with H 2 O, 0.5 mM SR3677 (SR) or H 2 O/SR3677 combined with 5 mM paraquat (PQ). Scale bars, 10 µm. d Quantification of the percentage of red-only mitochondrial area divided by the total non-background area, averaged across 0.8-µm z-stacks. Data are expressed as mean ± s.e.m ( n = 4 independent experiments). P -values were determined by one-tailed paired Student’s t -test, * P

    Journal: Nature Communications

    Article Title: ROCK inhibitors upregulate the neuroprotective Parkin-mediated mitophagy pathway

    doi: 10.1038/s41467-019-13781-3

    Figure Lengend Snippet: Targeting of mitochondria to lysosomes is increased by SR3677. a HeLa cells were co-transfected with Cerulean-Parkin and RG-OMP25. After 24 h, cells were pre-treated with either 0.5 µM SR3677 or DMSO for 2 h prior to induction of mitophagy with 10 µM CCCP treatment, in combination with E-64 and leupeptin. Red-only signal represents mitochondria localized to lysosomes, where GFP signal is quenched. Scale bars, 10 µm. b Quantification of the percentage of red-only mitochondrial area divided by the total non-background area ( n = 4 independent experiments). P -values were determined by one-tailed paired Student’s t -test. c Seven-day-old TH-GAL4 > UAS-mitoQC male flies were placed into vials containing the indicated treatments. Representative images of the dopaminergic neurons of TH-GAL4 > UAS-mitoQC flies following feeding on fly food supplemented with H 2 O, 0.5 mM SR3677 (SR) or H 2 O/SR3677 combined with 5 mM paraquat (PQ). Scale bars, 10 µm. d Quantification of the percentage of red-only mitochondrial area divided by the total non-background area, averaged across 0.8-µm z-stacks. Data are expressed as mean ± s.e.m ( n = 4 independent experiments). P -values were determined by one-tailed paired Student’s t -test, * P

    Article Snippet: Chemicals The following chemicals were used in our study: CCCP (Sigma, C2759), SR3677 (Tocris, 3667/10), Y27632 (Millipore, 688000), Y39983 (MedChemExpress, HY-10069), paraquat (Sigma, 36541), E-64 (BioShop, EEL640.1), leupeptin (BioShop, Leu001.50), geneticin (Gibco, 11811023), puromycin (BioShop, PUR333.10), retinoic acid (Calbiochem, 554720), brain-derived neurotrophic factor (Gibco, PHC7074) and chloroquine (Bioshop, CHL919).

    Techniques: Transfection, One-tailed Test

    Inhibition of CroP activity by aprotinin. (A) Aprotinin inhibits cleavage of the FRET substrate by purified CroP. FRET assays were performed with 10 nM CroP in PBS (black) or in the presence of EDTA (2.5 mM, green), PMSF (2.5 mM, purple), leupeptin (10

    Journal: Infection and Immunity

    Article Title: Inhibition of Outer Membrane Proteases of the Omptin Family by Aprotinin

    doi: 10.1128/IAI.00136-15

    Figure Lengend Snippet: Inhibition of CroP activity by aprotinin. (A) Aprotinin inhibits cleavage of the FRET substrate by purified CroP. FRET assays were performed with 10 nM CroP in PBS (black) or in the presence of EDTA (2.5 mM, green), PMSF (2.5 mM, purple), leupeptin (10

    Article Snippet: The protease inhibitors EDTA, leupeptin, pepstatin A, and aprotinin were from BioShop; phenylmethylsulfonyl fluoride (PMSF) was purchased from Sigma-Aldrich.

    Techniques: Inhibition, Activity Assay, Purification