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European Journal of Biochemistry Oct 19801. Phosphoprotein phosphatase IB is a form of rat liver phosphoprotein phosphatase, distinguished from the previously studied phosphoprotein phosphatase II [Tamura et...
1. Phosphoprotein phosphatase IB is a form of rat liver phosphoprotein phosphatase, distinguished from the previously studied phosphoprotein phosphatase II [Tamura et al. (1980) Eur. J. Biochem. 104, 347-355] by earlier elution from DEAE-cellulose, by higher molecular weight on gel filtration (260000) and by lower activity toward phosphorylase alpha. This enzyme was purified to apparent homogeneity by chromatography on DEAE-cellulose, aminohexyl--Sepharose-4B, histone--Sepharose-4B, protamine--Sepharose-4B and Sephadex G-200. 2. The molecular weight of purified phosphatase IB was 260000 by gel filtration and 185000 from S20,W and Stokes' radius. Using histone phosphatase activity as the reference for comparison, the phosphorylase phosphatase activity of purified phosphatase IB was only one-fifth that of phosphatase II. 3. Sodium dodecyl sulfate gel electrophoresis revealed that phosphatase IB contains three types of subunit, namely alpha, beta and gamma, whose molecular weights are 35000, 69000 and 58000, respectively. The alpha subunit is identical to the alpha subunit of phosphatase II. While the beta subunit is also identical or similar to the beta subunit of phoshatase II, the gamma subunit appears to be unique to phosphatase IB. 4. When purified phosphatase IB was treated with 2-mercaptoethanol at -20 degrees C, the enzyme was dissociated to release the catalytically active alpha subunit. Along with this dissociation, there was a 7.4-fold increase in phosphorylase phosphatase activity; but histone phosphatase activity increased only 1.6-fold. The possible functions of the gamma subunit are discussed in relation to this activation of enzyme.
Topics: Animals; Liver; Macromolecular Substances; Mercaptoethanol; Molecular Weight; Phosphoprotein Phosphatases; Rats
PubMed: 6254774
DOI: 10.1111/j.1432-1033.1980.tb06096.x -
The EMBO Journal Nov 2000Phosphoprotein phosphatase 2A (PP2A) is a major phosphoserine/threonine protein phosphatase in all eukaryotes. It has been isolated as a heterotrimeric holoenzyme...
Phosphoprotein phosphatase 2A (PP2A) is a major phosphoserine/threonine protein phosphatase in all eukaryotes. It has been isolated as a heterotrimeric holoenzyme composed of a 65 kDa A subunit, which serves as a scaffold for the association of the 36 kDa catalytic C subunit, and a variety of B subunits that control phosphatase specificity. The C subunit is reversibly methyl esterified by specific methyltransferase and methylesterase enzymes at a completely conserved C-terminal leucine residue. Here we show that methylation plays an essential role in promoting PP2A holoenzyme assembly and that demethylation has an opposing effect. Changes in methylation indirectly regulate PP2A phosphatase activity by controlling the binding of regulatory B subunits to AC dimers.
Topics: Amino Acid Sequence; Animals; Brain; Cattle; Enzyme Stability; Holoenzymes; In Vitro Techniques; Methylation; Phosphoprotein Phosphatases; Protein Methyltransferases; Protein Phosphatase 2; Protein Structure, Quaternary; Protein Subunits; Substrate Specificity
PubMed: 11060019
DOI: 10.1093/emboj/19.21.5682 -
Biochemical and Biophysical Research... Aug 1979
Topics: Animals; Cations, Divalent; Edetic Acid; Fluorides; Kinetics; Liver; Nuclear Envelope; Phosphoprotein Phosphatases; Rats
PubMed: 227377
DOI: 10.1016/0006-291x(79)92118-1 -
Methods in Enzymology 1998
Topics: Animals; Antibodies; Carrier Proteins; Cell Line; Chromatography, High Pressure Liquid; Cytoskeletal Proteins; Enzyme Inhibitors; Fluorescent Antibody Technique; Intracellular Signaling Peptides and Proteins; Keratins; Mutagenesis, Site-Directed; Organophosphorus Compounds; Peptide Mapping; Phosphoprotein Phosphatases; Phosphorylation; Precipitin Tests; Protein Kinase Inhibitors; Protein Kinases; Sequence Analysis; Solubility; Transfection; Vimentin
PubMed: 9751907
DOI: 10.1016/s0076-6879(98)98044-2 -
Current Opinion in Biotechnology Aug 1996In the past year, the three-dimensional structures of two serine/threonine phosphatases, protein phosphatase-1 and protein phosphatase-2b (calcineurin), have been... (Review)
Review
In the past year, the three-dimensional structures of two serine/threonine phosphatases, protein phosphatase-1 and protein phosphatase-2b (calcineurin), have been determined. The new information puts previous sequence comparisons and mutagenesis studies into a detailed structural perspective. The active-site structure and catalytic mechanism appear to be common to a variety of phosphoesterase enzymes.
Topics: Amino Acid Sequence; Calcineurin; Calmodulin-Binding Proteins; Crystallography, X-Ray; Models, Molecular; Molecular Sequence Data; Phosphoprotein Phosphatases; Protein Conformation; Protein Engineering; Protein Phosphatase 1; Recombinant Proteins; Serine; Threonine
PubMed: 8768897
DOI: 10.1016/s0958-1669(96)80114-5 -
Biochemical Pharmacology Feb 1999Protein phosphatases are responsible for keeping the signaling output of stimulus-activated protein kinases in check; but protein phosphatases are also themselves... (Review)
Review
Protein phosphatases are responsible for keeping the signaling output of stimulus-activated protein kinases in check; but protein phosphatases are also themselves targets and conveyors of biological signals. Among the major serine/threonine phosphatases, protein phosphatase 2A (PP2A) appears to play a privileged role in the regulation of cell growth and division. How PP2A is regulated is an intriguing question. This review will focus on the role of local protein-protein interactions in PP2A control. Work from a number of laboratories has shown that the catalytic activity, substrate specificity, and subcellular targeting of PP2A are regulated by a remarkably diverse range of regulatory subunits and enzyme inhibitors. On the pathological side, DNA tumor viruses subvert PP2A function by producing proteins that compete with specific regulatory subunits. By interfering with PP2A, these viral proteins can elicit changes in the activity of specific signal transduction pathways, such as the mitogen-activated protein kinase cascade. Recent data indicate that besides classical holoenzyme forms, a fraction of PP2A molecules are associated with novel partners implicated in signal transduction. PP2A biochemically and genetically interacts with the Tap42/alpha4 protein, which is part of a rapamycin-sensitive pathway that connects extracellular stimuli to the initiation of mRNA translation. PP2A also binds to CK2alpha, the catalytic subunit of CK2 (formerly casein kinase 2), and binding is sensitive to mitogenic signaling. The potent effect of quantitatively minor PP2A partners might be explained by a general requirement for docking interactions with substrates under intracellular conditions.
Topics: Animals; Phosphoprotein Phosphatases; Protein Kinases; Protein Phosphatase 2; Signal Transduction; Sirolimus; Substrate Specificity; Viral Proteins
PubMed: 9933020
DOI: 10.1016/s0006-2952(98)00245-7 -
Chemistry (Weinheim An Der Bergstrasse,... Jun 2018Protein phosphatases, while long overlooked, have recently become appreciated as drivers of both normal- and disease-associated signaling events. As a result, the... (Review)
Review
Protein phosphatases, while long overlooked, have recently become appreciated as drivers of both normal- and disease-associated signaling events. As a result, the spotlight is now turning torwards this enzyme family and efforts geared towards the development of modern chemical tools for studying these enzymes are well underway. This Minireview focuses on the evolution of chemical activity probes, both optical and covalent, for the study of protein phosphatases. Small-molecule probes, global monitoring of phosphatase activity through the use of covalent modifiers, and targeted fluorescence-based activity probes are discussed. We conclude with an overview of open questions in the field and highlight the potential impact of chemical tools for studying protein phosphatases.
Topics: Animals; Biomarkers; Fluorescent Dyes; Humans; Kinetics; Phosphoprotein Phosphatases; Signal Transduction
PubMed: 29338103
DOI: 10.1002/chem.201705194 -
The Journal of Biological Chemistry Mar 2016Cellular signaling through protein tyrosine phosphorylation is well established in mammalian cells. Although lacking the classic tyrosine kinases present in humans,...
Cellular signaling through protein tyrosine phosphorylation is well established in mammalian cells. Although lacking the classic tyrosine kinases present in humans, plants have a tyrosine phospho-proteome that rivals human cells. Here we report a novel plant tyrosine phosphatase from Arabidopsis thaliana (AtRLPH2) that, surprisingly, has the sequence hallmarks of a phospho-serine/threonine phosphatase belonging to the PPP family. Rhizobiales/Rhodobacterales/Rhodospirillaceae-like phosphatases (RLPHs) are conserved in plants and several other eukaryotes, but not in animals. We demonstrate that AtRLPH2 is localized to the plant cell cytosol, is resistant to the classic serine/threonine phosphatase inhibitors okadaic acid and microcystin, but is inhibited by the tyrosine phosphatase inhibitor orthovanadate and is particularly sensitive to inhibition by the adenylates, ATP and ADP. AtRLPH2 displays remarkable selectivity toward tyrosine-phosphorylated peptides versus serine/threonine phospho-peptides and readily dephosphorylates a classic tyrosine phosphatase protein substrate, suggesting that in vivo it is a tyrosine phosphatase. To date, only one other tyrosine phosphatase is known in plants; thus AtRLPH2 represents one of the missing pieces in the plant tyrosine phosphatase repertoire and supports the concept of protein tyrosine phosphorylation as a key regulatory event in plants.
Topics: Adenosine Diphosphate; Adenosine Triphosphate; Arabidopsis; Arabidopsis Proteins; Phosphoprotein Phosphatases; Phosphorylation; Protein Tyrosine Phosphatases
PubMed: 26742850
DOI: 10.1074/jbc.M115.683656 -
Molecular Cell Dec 2019Dynamic protein phosphorylation constitutes a fundamental regulatory mechanism in all organisms. Phosphoprotein phosphatase 4 (PP4) is a conserved and essential nuclear...
Dynamic protein phosphorylation constitutes a fundamental regulatory mechanism in all organisms. Phosphoprotein phosphatase 4 (PP4) is a conserved and essential nuclear serine and threonine phosphatase. Despite the importance of PP4, general principles of substrate selection are unknown, hampering the study of signal regulation by this phosphatase. Here, we identify and thoroughly characterize a general PP4 consensus-binding motif, the FxxP motif. X-ray crystallography studies reveal that FxxP motifs bind to a conserved pocket in the PP4 regulatory subunit PPP4R3. Systems-wide in silico searches integrated with proteomic analysis of PP4 interacting proteins allow us to identify numerous FxxP motifs in proteins controlling a range of fundamental cellular processes. We identify an FxxP motif in the cohesin release factor WAPL and show that this regulates WAPL phosphorylation status and is required for efficient cohesin release. Collectively our work uncovers basic principles of PP4 specificity with broad implications for understanding phosphorylation-mediated signaling in cells.
Topics: Amino Acid Sequence; Binding Sites; Conserved Sequence; Crystallography, X-Ray; HEK293 Cells; HeLa Cells; Humans; Phosphoprotein Phosphatases; Phosphorylation; Protein Binding; Substrate Specificity
PubMed: 31585692
DOI: 10.1016/j.molcel.2019.08.029 -
Methods in Molecular Biology (Clifton,... 2007This chapter introduces the main families of protein phosphatases encoded by the human genome and discusses their classification, overall structure, regulation, and... (Review)
Review
This chapter introduces the main families of protein phosphatases encoded by the human genome and discusses their classification, overall structure, regulation, and physiological functions in human health and diseases. The topics of redundancy, diversity, and dynamic expression in individual cell types are briefly introduced, and the importance of technological approaches to phosphatase research is emphasized.
Topics: Gene Expression Regulation, Enzymologic; Genome, Human; Humans; Mutation; Phosphoprotein Phosphatases; Protein Structure, Tertiary; RNA Interference
PubMed: 17200550
DOI: 10.1385/1-59745-267-X:9