Thermo Scientific™ Mass Spectrometry Grade Proteases

Improve sequence coverage and protein identification for mass spectrometry with high-quality, MS-grade proteases provided in stable liquid or lyophilized formats. Suitable for either in-solution or in-gel digestion workflows, each protease may be used individually or in combination to improve protein sequence coverage.

Catalog Number	Mfr. No.	Description	Quantity
PI90053	Thermo Scientific™ 90053	Asp-N Endoproteinase, MS Grade	2 μg
PI90056	Thermo Scientific™ 90056	Chymotrypsin Endoproteinase, MS Grade (TLCK-treated)	4 x 25 μg
PI84841	Thermo Scientific™ 84841	Digestion Indicator for Mass Spectrometry	10 μg
PI90054	Thermo Scientific™ 90054	Glu-C Endoproteinase, MS Grade	5 x 10 μg
PI90305	Thermo Scientific™ 90305	Trypsin Protease, MS Grade (Frozen)	1 x 100 μg
PI90059	Thermo Scientific™ 90059	Trypsin Protease, MS Grade	1 mg
PIA40007	Thermo Scientific™ A40007	Trypsin/Lys-C Protease Mix, MS Grade	20 μg
PIA41007	Thermo Scientific™ A41007	Trypsin/Lys-C Protease Mix, MS Grade	5 x 20 μg
PIA40009	Thermo Scientific™ A40009	Trypsin/Lys-C Protease Mix, MS Grade	100 μg
PI90051	Thermo Scientific™ 90051	Lys-C Endoproteinase, MS Grade	1 x 20 μg
FER90307	Thermo Scientific™ 90307	Lys-C Protease, MS Grade	100 μg
PI90057	Thermo Scientific™ 90057	Trypsin Protease, MS Grade	5 x 20 μg
PI90058	Thermo Scientific™ 90058	Trypsin Protease, MS Grade	5 x 100 μg

Description

Protein characterization, identification, and quantification by MS begins with efficient, reproducible protein digestion. Sequence specific proteases are utilized to cleave proteins into smaller fragments or peptides. Trypsin is the protease of choice for protein digestion. However, separate or sequential digestion with alternative proteases can improve individual protein sequence coverage, or generate unique peptide sequences for different MS applications.

• Exceptional selectivity—trypsin has >95% C-terminal lysine and arginine specificity; LysC has >90% C-terminal lysine cleavage specificity
• High purity—no detectable chymotrypsin activity (trypsin)
• Complete digestion—trypsin/LysC enzyme combination reduces tryptic missed cleavages
• Enhanced stability—trypsin chemically modified for reduced autolytic activity
• Convenient—enzymes provided in stable lyophilized or liquid formats

Trypsin Protease is a mass spectrometry (MS)-grade serine protease derived from porcine pancreatic extracts that specifically cleaves at the carboxyl side of lysine and arginine residues. The enzyme is TPCK-treated to eliminate chymotryptic activity and chemically modified by methylation, yielding a highly active and more stable form of the enzyme. Trypsin Protease can tolerate commonly used partially denaturing conditions, such as 0.1% SDS, 1 M urea, and 10% acetonitrile. Trypsin Protease is most active at pH 7 to 9 and can be reversibly inactivated at pH < 4. Trypsin can be purchased lyophilized in 1-mg 5-packs of 20 μg or 100 μg quantities and in solution at 100 μg.

Lys-C Protease is a mass spectrometry (MS)-grade serine protease isolated from Lysobacter enzymogenes. Lys-C Protease has high activity and specificity for lysine residues, resulting in larger peptides and less sample complexity than trypsin (i.e., fewer peptides). Unlike trypsin, Lys-C Protease can cleave lysines followed by prolines, making it ideal for sequential protein digestion followed by trypsin to decrease missed cleavages. These unique Lys-C Protease properties ensure high digestion efficiency when used alone or followed by tryptic digestion. Additionally, Lys-C prototypic peptides typically have higher charge states, making it an enzyme of choice for use with ETD fragmentation.

Lys-C Protease is commonly used in phosphopeptide enrichment workflows because it generates peptides with primary amines at both the N- and C-terminus, allowing the fragments to be double-labeled with amine-reactive isobaric tags. This results in enhanced peptide ionization and improved limits of quantitation since more fragment ions can be re-isolated during MS3 acquisition. This enzyme can be used for in-solution or in-gel digestion workflows to produce peptides for LC-MS/MS protein identification.

Efficient protein digestion can be completed in two hours at 37°C. Lys-C Protease remains active in highly denaturing conditions, such as 8 M urea, 2 M guanidine-HCl, 1% SDS, 2% CHAPS, and 40% acetonitrile, and functions well at pH 7–9 (maximal activity at pH 8). This lyophilized enzyme has a mass of 30 kDa and is stable for one year when stored at –20°C. This Lys-C enzyme is packaged lyophilized (20 μg or 100 μg quantities).

Trypsin/Lys-C Protease Mix is a lyophilized mixture of trypsin and LysC proteases that has been optimized to improve digestion efficiency of proteins. Although trypsin is routinely used for protein digestion, this protease alone is not sufficient to fully digest proteins at the carboxyl-end of lysine and arginine residues. Therefore, Lys-C protease is commonly combined with trypsin to sequentially digest proteins with fewer missed cleavages. Trypsin/Lys-C Protease Mix is provided in flexible formats of 20 μg, 5 x 20 μg, or 100 μg. Digestion can be completed in as little as 1.5–3 hours or up to overnight, depending on enzyme to protein ratio.

Chymotrypsin, GluC, Asp-N Proteases

• Increased sequence coverage—better protein characterization results from overlapping peptides with complementary chromatographic, ionization, and fragmentation properties
• High specific activity—each protease has excellent enzymatic specificity
• Stable—provided in a lyophilized format

Asp-N Protease is an MS-grade zinc metalloproteinase derived from a mutant strain of Pseudomonas fragi and requires a trace amount of zinc for activity. Asp-N Protease cleaves primarily at the amino side of aspartate and cysteic acid that results from cysteine oxidation. Cleavage can also occur at glutamic acid; however, the rate of cleavage at the glutamyl residue is significantly lower than the rate of cleavage at the aspartic acid residue. Asp-N Protease can efficiently digest protein in 2–20 hours at 37°C with activity greater than 20,000 units/mg protein and remains active under denaturing conditions, such as 1 M urea, 2 M guanidine·HCl, 0.1% SDS, 2% CHAPS, and 10% acetonitrile, with optimal activity at pH 6–8. This lyophilized enzyme has a mass of 27 kDa and is stable for one year when stored at -20°C.

Glu-C Protease, also referred to as V-8 protease, is an MS-grade serine protease isolated from Staphylococcus aureus. Glu-C Protease specifically cleaves the carboxyl side of glutamic acid residues in ammonium bicarbonate and ammonium acetate buffers, generating a limited number of peptide fragments. Cleavage can also occur at both glutamic and aspartic acid residues in phosphate buffers. Glu-C Protease can efficiently digest protein in 5–18 hours at 37°C with activity greater than 500 units/mg protein and remains active under denaturing conditions, such as 2 M urea, 1 M guanidine·HCl, 0.1% SDS, 2% CHAPS, and 20% acetonitrile. Glu-C Protease activity is optimal at pH 8. This lyophilized enzyme has a mass of 27 kDa and is stable for one year when stored at -20°C.

Chymotrypsin Protease is an MS-grade endoproteinase isolated from bovine pancreas that specifically cleaves at the carboxyl side of tyrosine, phenylalanine, tryptophan, and leucine. Two predominant forms of chymotrypsin protease, A and B, are found in equal amounts in bovine pancreas. They are similar proteins (80% homology) but have different proteolytic characteristics. Both forms of chymotrypsin protease are present in Thermo Scientific Chymotrypsin Protease. Since trypsin may co-purify with chymotrypsin protease derived from natural sources, Chymotrypsin Protease has been treated with TLCK to eliminate potential tryptic activity, improving digestion specificity. Chymotrypsin Protease can tolerate mild denaturing conditions, such as 0.1% SDS, 2 M urea, 2 M guanidine·HCl, 1% CHAPS, and 30% acetonitrile, with optimal activity at pH 7.5–8.5. This lyophilized enzyme has a mass of 25 kDa and is stable for one year when stored at -20°C.

Specifications

• For Use With (Equipment): Mass Spectrometer
• Product Line: Pierce™
• Detection Method: Mass Spectrometry
• Workflow Step: Protein Digestion