pkaPS: prediction of protein kinase A phosphorylation sites with the simplified kinase-substrate binding model

Neuberger, Georg; Schneider, Georg; Eisenhaber, Frank

doi:10.1186/1745-6150-2-1

Cited by 130 publications

(71 citation statements)

References 86 publications

(94 reference statements)

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“…Several experimental studies and various computational approaches have been employed extensively in search of substrates and prediction of specific phosphorylation sites both for mammalian (13,14) and yeast (21,15,19,42) systems, both for kinases in general (4,8,13,21) or for PKA in particular (14,15,19,42). Our results contribute to the phosphorylation dynamics in S. cerevisiae.…”

Section: Discussionmentioning

confidence: 68%

“…More recent reports on prediction of protein kinase A phosphorylation sites (13,42) confirmed and generalized these early predictions and established that the requirements for positively charged residues are highest for residues in positions P-2 and P-3 but can be detected as far as six to eight residues prior to the phosphorylated serine. Yeast PKA has been shown to have the same requirements of basic residues at positions P-2 and P-3 FIGURE 2.…”

Section: Resultsmentioning

confidence: 76%

“…No clear consensus sequence has been established for the C-terminal flank of the phosphorylatable Ser/ Thr, with exception of position ϩ1 with its large hydrophobic amino acid and a slight preference for small, flexible residues at positions ϩ2 and ϩ3 (42). Few data were available from the literature regarding Pϩ4 until recently, as this position was missing in the resolved three-dimensional structures (43).…”

Section: Resultsmentioning

confidence: 99%

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Characterization of Substrates That Have a Differential Effect on Saccharomyces cerevisiae Protein Kinase A Holoenzyme Activation

Galello

Portela

Moreno

et al. 2010

Journal of Biological Chemistry

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The specificity in phosphorylation by kinases is determined by the molecular recognition of the peptide target sequence. In Saccharomyces cerevisiae, the protein kinase A (PKA) specificity determinants are less studied than in mammalian PKA. The catalytic turnover numbers of the catalytic subunits isoforms Tpk1 and Tpk2 were determined, and both enzymes are shown to have the same value of 3 s ؊1 . We analyze the substrate behavior and sequence determinants around the phosphorylation site of three protein substrates, Pyk1, Pyk2, and Nth1. Nth1 protein is a better substrate than Pyk1 protein, and both are phosphorylated by either Tpk1 or Tpk2. Both enzymes also have the same selectivity toward the protein substrates and the peptides derived from them. The three substrates contain one or more Arg-Arg-X-Ser consensus motif, but not all of them are phosphorylated. The determinants for specificity were studied using the peptide arrays. Acidic residues in the position P؉1 or in the N-terminal flank are deleterious, and positive residues present beyond P-2 and P-3 favor the catalytic reaction. A bulky hydrophobic residue in position P؉1 is not critical. The best substrate has in position P؉4 an acidic residue, equivalent to the one in the inhibitory sequence of Bcy1, the yeast regulatory subunit of PKA. The substrate effect in the holoenzyme activation was analyzed, and we demonstrate that peptides and protein substrates sensitized the holoenzyme to activation by cAMP in different degrees, depending on their sequences. The results also suggest that protein substrates are better co-activators than peptide substrates.Protein phosphorylation is one of the most fundamental mechanisms for signal transduction. The specificity of signal transduction depends upon the ability of each kinase to precisely phosphorylate particular sites on specific substrate proteins. Protein kinase A (PKA) 3 is one of the best characterized members of this protein family (1). Phosphorylation of its substrate targets is known to be critical for regulating a multitude of cellular processes, including metabolism, gene transcription, ion flux, growth, and cell death (2). The specificity in phosphorylation is determined by at least two major factors: peptide specificity and substrate and kinase localization. The effectiveness of protein phosphorylation by protein kinases in general, including PKA, is believed to depend on the primary structure of the protein around the phosphorylation site; synthetic peptides have thus been used to define the consensus phosphorylation sequences for PKA (3-5). The basic consensus phosphorylation sites for this enzyme are Arg-Arg-X-(Ser/Thr), (Arg/Lys)-X-X-(Ser/Thr), and (Arg/Lys)-X-(Ser/ Thr) (3, 4, 6, 7).The molecular recognition of the peptide sequence surrounding the phosphorylated residue of a substrate usually is referred to as the "peptide specificity" of a protein kinase. The phosphorylated residue is conventionally named P0, the substrate residues immediately N-and C-terminal to the P0 position as PϪ1 and Pϩ1, r...

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Section: Discussionmentioning

confidence: 68%

Section: Resultsmentioning

confidence: 76%

Section: Resultsmentioning

confidence: 99%

See 1 more Smart Citation

Characterization of Substrates That Have a Differential Effect on Saccharomyces cerevisiae Protein Kinase A Holoenzyme Activation

Galello

Portela

Moreno

et al. 2010

Journal of Biological Chemistry

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“…To understand the role of S253 phosphorylation in regulating the chromosome binding function of the HPV8 E2 protein, we proceeded to first identify the cellular kinase responsible for S253 phosphorylation. Using the pkaPS predictor program, which predicts PKA phosphorylation sites on the basis of a simplified kinase substrate binding model (27), S253 was predicted to be a strong PKA substrate site (data not shown). Out of the 59 serine residues in the HPV8 E2 protein, serine 253 had the highest score.…”

Section: Resultsmentioning

confidence: 99%

Phosphorylation Regulates Binding of the Human Papillomavirus Type 8 E2 Protein to Host Chromosomes

Sekhar

McBride

2012

J Virol

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The papillomavirus E2 proteins are indispensable for the viral life cycle, and their functions are subject to tight regulation. The E2 proteins undergo posttranslational modifications that regulate their properties and roles in viral transcription, replication, and genome maintenance. During persistent infection, the E2 proteins from many papillomaviruses act as molecular bridges that tether the viral genomes to host chromosomes to retain them within the host nucleus and to partition them to daughter cells. The betapapillomavirus E2 proteins bind to pericentromeric regions of host mitotic chromosomes, including the ribosomal DNA loci. We recently reported that two residues (arginine 250 and serine 253) within the chromosome binding region of the human papillomavirus type 8 (HPV8) E2 protein are required for this binding. In this study, we show that serine 253 is phosphorylated, most likely by protein kinase A, and this modulates the interaction of the E2 protein with cellular chromatin. Furthermore, we show that this phosphorylation occurs in S phase, increases the half-life of the E2 protein, and promotes chromatin binding from S phase through mitosis.

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“…Prediction software (pkaPS) from the IMP-IMBA Bioinformatics group [50] suggested that the recognition motif around S811 of NCC SV shows high similarity to a protein kinase A (PKA) consensus site. Interestingly, several hormonal systems have been shown to activate the cAMP-PKA pathway [51,52].…”

Section: Functional Role Of Nccsvmentioning

confidence: 99%

Role of the alternative splice variant of NCC in blood pressure control

2018

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The renal thiazide-sensitive sodium-chloride cotransporter (NCC), located in the distal convoluted tubule (DCT) of the kidney, plays an important role in blood pressure regulation by fine-tuning sodium excretion. The human SLC12A3 gene, encoding NCC, gives rise to three isoforms, of which only the third isoform (NCC3) has been extensively investigated so far. However, recent studies unraveled the importance of the isoforms 1 and 2, collectively referred to as NCC splice variant (NCCSV), in several (patho)physiological conditions. In the human kidney, NCCSV localizes to the apical membrane of the DCT and could constitute a functional route for renal sodium-chloride reabsorption. Analysis of urinary extracellular vesicles (uEVs), a non-invasive method for measuring renal responses, demonstrated that NCCSV abundance changes in response to acute water loading and correlates with patients’ thiazide responsiveness. Furthermore, a novel phosphorylation site at serine 811 (S811), exclusively present in NCCSV, was shown to play an instrumental role in NCCSV as well as NCC3 function. This review aims to summarize these new insights of NCCSV function in humans that broadens the understanding on NCC regulation in blood pressure control.

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pkaPS: prediction of protein kinase A phosphorylation sites with the simplified kinase-substrate binding model

Cited by 130 publications

References 86 publications

Characterization of Substrates That Have a Differential Effect on Saccharomyces cerevisiae Protein Kinase A Holoenzyme Activation

Characterization of Substrates That Have a Differential Effect on Saccharomyces cerevisiae Protein Kinase A Holoenzyme Activation

Phosphorylation Regulates Binding of the Human Papillomavirus Type 8 E2 Protein to Host Chromosomes

Role of the alternative splice variant of NCC in blood pressure control

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