Table

Table 1. Inhibitor cocktails used to control proteolysis during protein isolation

Tissue type Protease inhibitors (working concentration) Target protease typea Stock solutionb
Animal tissues AEBSF (0.2 mM) Serine AEBSF: 20 mM in methanol
or
DCI (0.1 mM) DCI: 10 mM in DMSO
or
PMSF (0.2 mM) PMSF: 200 mM in ethanol or isopropanol
Benzamidine (1 mM) Serine 100 mM
Leupeptin (10 μg/mL) Serine/cysteine 1 mg/mL
Pepstatin (10 μg/mL) Aspartic 5 mg/mL in methanol
Aprotinin (1 μg/mL) Serine 0.1 mg/mL
EDTA or EGTAc (1 mM) Metallo 100 mM
Plant tissues AEBSF (0.2 mM) Serine AEBSF: 20 mM in methanol
or
DCI (0.1 mM) DCI: 10 mM in DMSO
or
PMSF (0.2 mM) PMSF: 200 mM in ethanol or isopropanol
Chymostatin (20 μg/mL) Serine/cysteine 1 mg/mL in DMSO
EDTA or EGTAc (1 mM) Metallo 100 mM
Yeasts and fungi AEBSF (0.2 mM) Serine AEBSF: 20 mM in methanoll
or
DCI (0.1 mM) DCI: 10 mM in DMSO
or
PMSF (0.2 mM) PMSF: 200 mM in ethanol or isopropano
Pepstatin (15 μg/mL) Aspartic 5 mg/mL in methanol
1,10-Phenanthroline (5 mM) Metallo 1 M in ethanol
Bacteria AEBSF (0.2 mM) Serine AEBSF: 20 mM in methanol
or
DCI (0.1 mM) DCI: 10 mM in DMSO
or
PMSF (0.2 mM) PMSF: 200 mM in ethanol or isopropanol
EDTA or EGTAc (1 mM) Metallo 100 mM
Adapted from North (1989).
Abbreviations: AEBSF: 4-(2-aminoethyl)-benzenesulfonyl fluoride; DCI: 3,4-dichloroisocoumarin; DMSO: dimethyl sulfoxide; EDTA: ethylenediamine tetraacetic acid; EGTA: ethylene glycol bis(β-aminoethyl ether) N,N,N′,N′-tetraacetic acid; PMSF: phenylmethylsulfonyl fluoride.
Mr values of inhibitors: 1,10-phenanthroline: 198; AEBSF: 240; aprotinin: 6500; benzamidine hydrochloride: 157; chymostatin: 605; DCI: 215; EDTA (disodium salt, dihydrate): 372; leupeptin: 427; pepstatin: 686; PMSF: 174.
aFor a review of proteolytic enzymes, see Neurath (1989) and Perlmann and Lorand (1970).
bAqueous solution unless otherwise indicated.
cAn efficient chelator of divalent metal cations other than Mg2+ (for which it has a 103-fold lower affinity) (Gegenheimer 1990).
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  1. doi:10.1101/pdb.tab195466 Cold Spring Harb Protoc 2010: pdb.tab195466-

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