Protease substrates

X-GAL (BCIG) is a widely used chromogenic substrate for β-galactosidase, which cleaves X-GAL to produce an insoluble blue compound.

4-MUNANA is a fluorescent substrate employed in neuraminidase activity assays and serves as a neuraminidase substrate analogue.

ABTS diammonium salt (AzBTS-(NH4)2) is used as a substrate for horseradish peroxidase (HRP) conjugate.

Boc-Gln-Ala-Arg-AMC hydrochloride is a fluorescent compound used as a substrate for trypsin and for assessing the protein hydrolysis activity of TMPRSS2.

BCIP (BCIP p-toluidine salt) (BCIP p-toluidine salt) is an artificial chromogenic substrate. It is used for the sensitive colorimetric detection of alkaline phosphatase activity.

Z-IETD-AFC is a fluorescent substrate for caspase-8 and granzyme B. It can be used to quantify cysteinyl aspartate-8 or granzyme B activity.

Dansylcadaverine (Monodansyl cadaverine) is a fluorescent probe.

4-Methylumbelliferyl-β-D-glucuronide hydrate (MUG) is a fluorogenic substrate (λex=362 nm, λem=445 nm).

BCDA (5-bromo-4-chloroindoxyl acetate) is a chromogenic substrate utilized to detect esterase activity.

Msr-Ratio (Msr-green) (Msr-green) is a ratiometric fluorescent probe specifically designed for the detection of methionine sulfoxide reductase (MSR) activity. It has an excitation wavelength of 375 nm and an emission wavelength of 550 nm. This probe offers the ability to monitor the enzyme activity both in vitro and in live cells, making it a valuable tool for studying MSR function.

Mca-PLAC(p-OMeBz)-WAR(Dpa)-NH2 is a fluorogenic substrate for matrix metalloproteinase-14 (MMP-14). Upon cleavage by MMP-14, 7-methoxycoumarin-4-acetyl (Mca) is released and its fluorescence can be used to quantify MMP activity. Mca displays excitation emission maxima of 328 420 nm, respectively.

Z-Arg-Arg-4MβNA triacetate serves as a specific substrate for cathepsin B, producing the fluorescent product 4MβNA, which has an excitation wavelength (λ ex) of 355 nm and an emission wavelength (λ em) of 430 nm [1] [2].

Mca-PLGL-Dpa-AR-NH2 is a fluorescent peptide MMP substrate [1] .

Mca-Ala-Pro-Lys(Dnp)-OH, a specific quenched fluorogenic substrate for ACE2, enables the detection of ACE2 activity in various tissues, including urinary, heart, and lung.

Fast Green free acid is a dye that resists acid once applied.

TFMU-ADPr serves as a versatile substrate for assessing poly(ADP-ribose) glycohydrolase (PARG) activity, enabling direct measurement of total PAR hydrolase activity through the liberation of a fluorophore. Exhibiting exceptional reactivity, generality, stability, and usability, TFMU-ADPr is an effective tool for evaluating small-molecule inhibitors and investigating the regulation of ADP-ribosyl catabolic enzymes in vitro [1].

DABCYL-Glu-Arg-Nle-Phe-Leu-Ser-Phe-Pro-EDANS is a fluorescent dye used as a fluorogenic substrate for aspartyl proteinase from the human malaria parasite [1]. It should be stored protected from light.

D-Ala-Lys-AMCA, a known substrate of proton-coupled oligopeptide transporter 1 (PEPT1), emits blue fluorescence and may be transported into Caco-2 cells and liver cancer cells.

4-Methylumbelliferyl phosphate disodium (4-MUP), an anionic organophosphate, serves as a fluorogenic substrate for acid and alkaline phosphatases. Additionally, this compound acts as a simulant for nerve agents [1] [2] [3].

Bz-FVR-AMC is a fluorogenic substrate utilized for procathepsin, demonstrating a catalytic efficiency (k_cat K_m) of 1070 mM^-1 s^-1. Notably, elevated levels of Bz-FVR-AMC can inhibit the substrate [1][2]. For storage, it is essential to protect this compound from light exposure.

Thymolphthalein monophosphate disodium hydrate serves as a chromogenic substrate for the detection of acid phosphatase and alkaline phosphatase. During the enzymatic reaction, thymolphthalein is liberated which elevates the medium's pH, facilitating easy detection through color production and halts hydrolysis. Additionally, this compound is utilized for the specific identification of prostatic phosphatase in serum [1] [2].

Mca-YVADAP-Lys(Dnp)-OH serves as a fluorogenic substrate specifically for caspase-1 and angiotensin-converting enzyme 2 (ACE2) [1].

Z-DEVD-AMC is a selective substrate for caspase-3, measurable through fluorescence spectrometry, and serves as a fluorescence reference standard for AMC-based enzyme substrates, including those for caspase [1].

Dnp-PLGLWA-DArg-NH2 TFA, a fluorogenic substrate specifically for MMP-1 and MMP-9, facilitates the quantification of MMP activity at an excitation wavelength of 280 nm and an emission wavelength of 360 nm [1] [2].

L-Leu-AMC is a fluorogenic substrate for leucine aminopeptidase.1Upon enzymatic cleavage by leucine aminopeptidase, AMC is released and its fluorescence can be used to quantify leucine aminopeptidase activity. AMC displays excitation emission maxima of 340-360 440-460 nm, respectively. 1.Izquierdo, M., Lin, D., O'Neill, S., et al.Development of a high-throughput screening assay to identify inhibitors of the major M17-leucyl aminopeptidase from Trypanosoma cruzi using rapidfire mass spectrometrySLAS Discov.25(9)1064-1071(2020)

CUG is a fluorogenic substrate (λex=386, λem=445 nm, ε=32K).

Z-(L-Arg)-AMC is a fluorogenic substrate for trypsin, cathepsin B, and cathepsin H.1,2Upon enzymatic cleavage by trypsin, cathepsin B, or cathepsin H, 7-amino-4-methylcoumarin (AMC) is released and its fluorescence can be used to quantify trypsin, cathepsin B, and cathepsin H activity. AMC displays excitation emission maxima of 340-360 440-460 nm, respectively. 1.Zimmerman, M., Ashe, B., Yurewicz, E.C., et al.Sensitive assays for trypsin, elastase, and chymotrypsin using new fluorogenic substratesAnal. Biochem.78(1)47-51(1977) 2.Brindley, P.J., Kalinna, B.H., Dalton, J.P., et al.Proteolytic degradation of host hemoglobin by schistosomesMol. Biochem. Parasitol.89(1)1-9(1997)

Ac-EEVVAC-pNA serves as a chromogenic substrate specifically designed for the continuous spectrophotometric assay of the HCV NS3 protease. Its sequence, EEVVAC, originates from the 5A-5B cleavage junction within the HCV polyprotein [1].

Ac-ANW-AMC is a fluorogenic substrate for the β5i LMP7 subunit of the 20S immunoproteasome. [1] Upon cleavage, 7-amino-4-methylcoumarin (AMC) is released, and its fluorescence quantifies the activity of the β5i LMP7 subunit. AMC displays excitation emission maxima of 340-360 440-460 nm, respectively.

Proteinase 3 (PR3, myeloblastin) is a polymorphonuclear leukocyte serine proteinase that degrades matrix proteins including fibronectin, laminin, vitronectin, and collagen type IV to generate antimicrobial peptides. Neutrophil elastase is a serine proteinase that is secreted by neutrophils during inflammation to destroy pathogens. Evaluating these enzymes is helpful to understanding inflammatory autoimmune processes. MeOSuc-AAPV-pNA is a highly sensitive peptide substrate that is hydrolyzed by both human and mouse neutrophil elastase and PR3, but not cathepsin G or chymotrypsin. Enzyme activity can be quantified by colorimetric detection of free p-nitroanilide at 405 nm.

RH 421 is an amphiphilic styryl dye.

Fluorescein Di-β-D-Glucuronide, a fluorescent probe, enables non-invasive imaging of intestinal bacterial β-glucuronidase activity in nude mice. Its utilization for both in vitro and in vivo imaging provides valuable insights into bacterial β-glucuronidase activity, thereby enhancing pharmacodynamic research on specific inhibitors in animal models [1].

Mca-P-Cha-G-Nva-HA-Dap(DNP)-NH2 is a fluorogenic substrate used for quantifying the activity of matrix metalloproteinase-1 (MMP-1), MMP-3, and MMP-26, thus facilitating the measurement of these enzymes' activity [1] [2].

4-Methylumbelliferyl-β-D-galactoside (MUG) is a fluorogenic substrate for β-galactosidase (β-gal).1Hydrolysis of MUG by β-gal releases the fluorescent moiety 4-MU, which displays a pH-dependent excitation maximum of 330, 370, and 385 nm at pH 4.6, 7.4, and 10.4, respectively, and an emission maximum between 445-454 nm.2MUG has been used to detect β-gal activity in intact bacteria, yeast, and mammalian cells.1 1.Vidal-Aroca, F., Giannattasio, M., Brunelli, E., et al.One-step high-throughput assay for quantitative detection of β-galactosidase activity in intact Gram-negative bacteria, yeast, and mammalian cellsBiotechniques40(4)433-434(2006) 2.Zhi, H., Wang, J., Wang, S., et al.Fluorescent properties of hymecromone and fluorimetric analysis of hymecromone in compound dantong capsuleJ. Spectrosc.1(1)147128(2013)