Description:
Human recombinant protein phosphatase inhibitor-1 is produced from E. coli., purified from freshly extracted E. coli lysates, and purified to homogeneity using methods developed by LAE Biotechnology Co., LTD. The final fraction of protein phosphatase inhibitor-1 exhibits a single polypeptide band of 27 kDa. Protein phosphatase inhibitor-1 can be phosphorylated by cAMP-dependent protein kinase. Protein phosphatase inhibitor-1 after phosorylation on Thr-35 becomes a potent inhibitor of type 1 protein phosphatase (1, 2). Phospho-protein phosphatase inhibitor-1 specifically inhibits the catalytic subunit of type 1 protein phosphatases (PP1) at nanomolar concentrations (IC₅₀ ~ 2 nM). Protein phosphatase inhibitor-1 was prepared by phosphorylation on Thr-35 by cAMP-dependent protein kinase.

Quality of Phospho-Protein Phosphatase Inhibitor-1:

1X PP1 Reaction Buffer:
50 mM Tris-HCl, 5 mM dithiothreitol, 0.1 mM EDTA, pH 7.0

Reaction Conditions:
1X PP1 Reaction Buffer supplemented with 1 mM MnCl₂ and 5 mM Caffeine
Reaction should be performed at 30 C.

Supply:
Protein phosphatase inhibitor-1 is supplied in lyophilized form at the dose of 100 ug per vail (Catalog # P025) and at the dose of 50 ug per vail (Catalog # P026).

Reconstitution:
Lyophilized protein phosphatase inhibitor-1 (catalog # P025) should be reconstituted in 100 ul of 50 mM Tris-HCl, pH 7.0, while lyophilized protein phosphatase inhibitor-1 (catalog # P026) should be reconstituted in 50 ul of 50 mM Tris-HCl, pH 7.0, to make a final concentration of 1 mg/ml.

Shipping/Storage:
The protein phosphatase inhibitor-1 is shipped on ice and must be stored at -20 C or lower.

Legal consideration: FOR RESEARCH USE ONLY

Notes:
1. Supplement with 1 mM MnCl₂ and 5 mM caffeine (required only if phosphorylase a is the substrate).

2. Protein phosphatase inhibitor-1 has been purified to > 95% homogeneity as determined by SDS-PAGE and Coomassie Blue staining.

3. Recommended long term storage -70 C; avoid repeated freeze/thaw cycles.

Quality Assurance Statement:
Protein phosphatase inhibitor-1 contains no detectable protease activity. Tests for phosphatase activity showed no detectable activity.

References:
1. Connor, J. H. (1998) Inhibitor-1 interaction domain that mediates the inhibition of protein phosphatase-1. J. Biol. Chem. 273, 27716.

2. Connor, J. H. (1999) Importance of the beta12-beta13 loop in protein phosphatase-1 catalytic subunit for inhibition by toxins and mammalian protein inhibitors. J. Biol. Chem. 274, 22366.