Protease and phosphatase inhibitors are essential components of most cell lysis and protein extraction procedures. These inhibitors block or inactivate endogenous proteolytic and phospholytic enzymes that are released from subcellular compartments during cells lysis and would otherwise degrade proteins of interest and their activation states.

Proteases and Phosphatases

Proteases and phosphatases are important enzymes in a variety of biochemical pathways in living cells. Proteases are required for many cellular functions, including cellular repair and the digestion of extracellular material. Phosphatases play a key role in regulating signal transduction events in eukaryotic cells. Protein kinases transfer a phosphate from ATP to a serine, threonine or tyrosine residue in a protein; phosphatases remove the phosphoryl group. Phosphorylation is the most common post-translational modification on proteins, with approximately 80% occurring on serine, 20% on threonine and 0.1-1% on tyrosine residues.

All living organisms contain proteolytic enzymes (proteases and peptidases). In whole cells, protease and phosphatase activities are tightly regulated by compartmentalization or inhibitors to prevent indiscriminate damage to cellular proteins and to maintain proper function of signaling pathways. Cell lysis disturbs the carefully controlled cellular environment, allowing proteases and phosphatases to become unregulated. The usual consequence of this unregulated state is reduced recovery of total protein and biologically meaningless representation of protein activities (i.e., phosphorylation status).

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Cell Lysis Solutions
Protease Selection Guide

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MS-grade Endoproteinases
Protease Assay Kits
Pierce Cell Lysis Reagents

Inhibitors of Proteases and Phosphatases

Protease inhibitors are biological or chemical compounds that function by reversibly or irreversibly binding to the protease. Most known proteases belong to one of four evolutionarily distinct enzyme families based on the functional groups involved in cleavage of the peptide bond. Known phosphatases are specific for cleavage of either serine-threonine or tyrosine phosphate groups. Thus, while numerous compounds have been identified and used to inactivate or block these enzymes, no single chemical is effective for all types of proteases and phosphatases (Table).

Rather, most researchers prepare or use a mixture or "cocktail" of several different inhibitor compounds to ensure that protein extracts do not degrade before analysis for targets of interest. Proteases inhibitors are nearly always needed, while phosphatase inhibitors are required only when phosphorylation states (activation states) are being investigated. Particular research experiments may necessitate the use of single inhibitors or customized mixtures, but most protein work is best served by using a suitable protease inhibitor cocktail (see additional discussion below table).

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Protease Inhibitors
(individual compounds)

Inhibitor Cocktails

We offer a variety of both individual protease inhibitors and broad-spectrum protease and phosphatase inhibitor cocktails to accommodate specific and general needs in cell lysis and protein extraction methods. These inhibitors are suitable for the protection of proteins during their extraction from cultured cells, animal tissues, plant tissues, yeast or bacteria.

Halt Protease Inhibitor Cocktails target serine-, cysteine- and aspartic acid proteases and aminopeptidases. Metalloproteases are inhibited by the optional addition of EDTA. Halt Phosphatase Inhibitor Cocktails contain chemical compounds that target serine/threonine and tyrosine phosphatases. For convenience, components of the separate protease and phosphatase cocktails have been combined into an all-in-one solution, Halt Protease and Phosphatase Inhibitor Cocktail. The latter prevents protein degradation and preserves phosphorylation simultaneously, providing protection that is similar to the individual cocktails. All of the Halt Inhibitor Cocktails are provided in a liquid format for easy dispensing and are compatible with Thermo Scientific Pierce Protein Extraction Reagents..

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Halt Protease Inhibitor Cocktails
Halt Phosphatase Inhibitor Cocktails
Halt Protease and Phosphatase Inhibitor Cocktails