DOI: 10.1073/pnas.100460897. Ser <sup>67</sup> -phosphorylated inhibitor 1 is a potent protein phosphatase 1 inhibitor

Ser 67 -phosphorylated inhibitor 1 is a potent protein phosphatase 1 inhibitor

10.1073/pnas.100460897

Crossref journal-article

Proceedings of the National Academy of Sciences

Proceedings of the National Academy of Sciences (341)

Abstract

Inhibitor 1 (I-1) is a protein inhibitor of protein phosphatase 1 (PP1), a major eukaryotic Ser/Thr phosphatase. Nonphosphorylated I-1 is inactive, whereas phosphorylated I-1 is a potent PP1 inhibitor. I-1 is phosphorylated in vivo on Thr 35 and Ser 67 . Thr 35 is phosphorylated by cAMP-dependent protein kinase (A kinase), and Thr 35 -phosphorylated I-1 inhibits PP1. Until now the kinase that phosphorylates Ser 67 had not been identified and the physiological role of Ser 67 phosphorylation was unknown. In this study we detected a high level of kinase activity in brain extract when a glutathione S -transferase (GST) fusion I-1 mutant containing an Ala substituted for Thr 35 [GST-I-1(T35A)] was used as the substrate. GST-I-1(T35A) kinase and neuronal cdc2-like protein kinase (NCLK) in the brain extract could not be separated from each other by a series of sequential chromatographies. GST-I-1(T35A) kinase immunoprecipitated with anti-NCLK antibody from kinase-active column fractions. Purified NCLK-phosphorylated GST-I-1(T35A) and I-1 (0.7 mole of phosphate per mole of I-1). HPLC phosphopeptide mapping, amino acid sequencing, and site-directed mutagenesis determined that NCLK phosphorylates Ser 67 of I-1. NCLK-phosphorylated I-1 and I-1(T35A) inhibited PP1 with IC 50 values ≈9.5 and 13.8 nM, respectively. When compared, A kinase-phosphorylated I-1 was only ≈1.2 times more inhibitory than NCLK-phosphorylated I-1. Our data indicate that NCLK is a potential in vivo I-1 kinase and that Thr 35 and Ser 67 phosphorylation independently activate I-1.

Bibliography

Huang, K., & Paudel, H. K. (2000). Ser 67 -phosphorylated inhibitor 1 is a potent protein phosphatase 1 inhibitor. Proceedings of the National Academy of Sciences, 97(11), 5824â5829.

Authors 2

Ke-xue Huang (first)
Hemant K. Paudel (additional)

References 30 Referenced 42

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Dates

Type	When
Created	23 years, 1 month ago (July 26, 2002, 10:41 a.m.)
Deposited	3 years, 4 months ago (April 12, 2022, 6:46 p.m.)
Indexed	1 year ago (Aug. 4, 2024, 1:18 a.m.)
Issued	25 years, 3 months ago (May 16, 2000)
Published	25 years, 3 months ago (May 16, 2000)
Published Online	25 years, 3 months ago (May 16, 2000)
Published Print	25 years, 3 months ago (May 23, 2000)

Funders 0

None

BibTeX

@article{Huang_2000, title={Ser 67 -phosphorylated inhibitor 1 is a potent protein phosphatase 1 inhibitor}, volume={97}, ISSN={1091-6490}, url={http://dx.doi.org/10.1073/pnas.100460897}, DOI={10.1073/pnas.100460897}, number={11}, journal={Proceedings of the National Academy of Sciences}, publisher={Proceedings of the National Academy of Sciences}, author={Huang, Ke-xue and Paudel, Hemant K.}, year={2000}, month=may, pages={5824–5829} }

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