Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.11.13 (protein kinase C)
 49,245 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Three monoclonal antibodies were prepared against rat brain soluble protein kinase C. Two of the antibodies, CKI-97 (IgG2b subclass) and CKII-90 (IgG1 subclass), showed weak binding to native protein kinase C. The third antibody, CKI-33 (IgG2b subclass), showed no binding. However, the mixture of CKI-97, CKII-90, and CKI-33 exhibited much stronger binding activity to this protein kinase than any of the antibodies alone. Although none of these antibodies showed protein kinase C-neutralizing activity, Western blot analysis indicated that these antibodies reacted specifically with protein kinase C, presumably its subspecies, that is present predominantly in nervous tissues. Immunocytochemical studies shows that these antibodies can be used for identification of this enzyme in nervous tissues. In rat Purkinje cells, the immunoreactive material was present throughout the cytoplasm, including dendrites and axons, but was poorly represented in the cell nucleus. In cerebellum, the localization of protein kinase C appears to be very similar to that of cGMP-dependent protein kinase.
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PMID:Monoclonal antibodies against rat brain protein kinase C and their application to immunocytochemistry in nervous tissues. 355 49

Rat liver cytosolic CKII shows heterogeneity resulting from association of the alpha/alpha'-subunits with the beta-subunit or with a phosphorylatable protein of 49 kDa (pp49). Preparations of pp49 were resolved into several spots by two dimensional analysis which might be derived from different degrees of phosphorylation. pp49 was phosphorylated in vitro by purified rat liver CKII and to a lower extent by purified rat brain protein kinase C. In all cases, phosphorylation of pp49 occurred exclusively on Ser. Phosphopeptide maps of phosphorylated pp49 confirmed that the phosphorylation by CKII or PKC takes place in different sites. Prior phosphorylation of pp49 by protein kinase C had no significant influence on the increase of the Km value for beta-casein of CKII, caused by pp49. A tryptic peptide from pp49 has been recently sequenced and antibodies against it had been raised. The antibodies were able to recognize pp49 in rat liver extracts as well as in HL-60 extracts what leads us to presume that this kind of interaction might exist in other species and tissues.
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PMID:Protein kinase CKII: possible regulation by interaction with protein substrates. 773 19

Protein kinases exhibit substrate specificities that are often primarily determined by the amino acids around the phosphorylation sites. Peptides corresponding to protein kinase C phosphorylation sites in several different proteins were synthesized on SPOTs membrane which has recently been found to be applicable for studies of protein kinase specificity. After phosphorylation with protein kinase C, we chose the best phosphorylated peptides for the investigation of the importance of amino acids immediately adjacent to the phosphorylation site. The selectivity of the best protein kinase C substrates from this study was analysed with protein kinases A, CK1 and CK2. According to these tests, the most favourable characteristics of SPOTs-membrane-associated peptides were demonstrated by peptide KRAKRKTAKKR. Kinetic analysis of peptide phosphorylation with protein kinase C revealed an apparent Km of 0.49 +/- 0.13 microM and Vmax of 10.0 +/- 0.5 nmol/min per mg with soluble peptide KRAKRKTAKKR. In addition, we assayed several other soluble peptides commonly used as protein kinase C substrates. Peptide KRAKRKTAKKR showed the lowest Km and the highest Vmax/Km value in comparison with peptides FKKSFKL, pEKRPSQRSKYL and KRAKRKTTKKR. Furthermore, of the peptides tested, KRAKRKTAKKR was the most selective substrate for protein kinase C. The favourable kinetic parameters combined with the selectivity should make the KRAKRKTAKKR peptide useful as a substrate for protein kinase C in the assays of both purified enzyme and in crude cell extracts.
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PMID:A potent and highly selective peptide substrate for protein kinase C assay. 906 63

IRF2 is a transcription factor, possessing oncogenic potential, responsible for both the repression of growth-inhibiting genes (interferon) and the activation of cell cycle-regulated genes (histone H4). Surprisingly little is known about the post-translational modification of this factor. In this study, we analyze the phosphorylation of IRF2 both in vivo and in vitro. Immunoprecipitation of HA-tagged IRF2 expressed in 32P-phosphate labelled COS-7 cells demonstrates that IRF2 is phosphorylated in vivo. Amino acid sequence analysis reveals that several potential phosphorylation sites exist for a variety of serine/threonine protein kinases, including those of the mitogen activated protein (MAP) kinase family. Using a battery of these protein kinases we show that recombinant IRF2 is a substrate for protein kinase A (PKA), protein kinase C (PKC), and casein kinase II (CK2) in vitro. However, other serine/threonine protein kinases, including the MAP kinases JNK1, p38, and ERK2, do not phosphorylate IRF2. Two-dimensional phosphopeptide mapping of the sites phosphorylated by PKA, PKC, and CKII in vitro demonstrates that these enzymes are capable of phosphorylating IRF2 at multiple distinct sites. Phosphoaminoacid analysis of HA-tagged IRF2 immunoprecipitated from an asynchronous population of proliferating, metabolically phosphate-labelled cells indicates that this protein is phosphorylated exclusively upon serine residues in vivo. These results suggest that the oncogenic protein IRF2 may be regulated via multiple pathways during cellular growth.
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PMID:Phosphorylation of the oncogenic transcription factor interferon regulatory factor 2 (IRF2) in vitro and in vivo. 921 19

It was our aim in this study to investigate the possibility that the third component of complement (C3) is phosphorylated during synthesis and secretion in U937 cells. Labelling of U937 cells with [32P]Pi, followed by immunoprecipitation of C3 from cell lysates and culture supernatants at different time points, showed that C3 was phosphorylated intracellularly immediately before release into the medium, which initiated cleavage of the protein into an iC3b-like fragment. Stimulation of CD11b/CD18 increased phosphorylation 7-fold, from a basal level of 2%. The phosphorylation sites in C3 did not resemble those described previously for casein kinase (CK) 1, cAMP-dependent protein kinase A or calcium- and phospholipid-dependent protein kinase C. Instead, protein kinase CK2 was suggested inasmuch as: (1) CK2 was detected both on the cell surface and on shed microparticles; (2) phosphorylation of purified C3 by microparticles was abolished by a monoclonal antibody, anti-CK2; (3) the [32P]Pi tag of both phosphorylated C3 (secreted from U937 cells) and of microparticle-phosphorylated C3, which was cleaved either by membrane proteases or by leucocyte elastase, was found in a 40 and a 70 kDa polypeptide; (4) both secreted C3 and C3 phosphorylated in vitro were much more susceptible to cleavage by proteases. Generation of C3 fragments provides a means by which U937 cells can stimulate nearby cells which are expressing complement receptors. The present study demonstrates that the cleavage of C3 is controlled by an intracellular phosphorylation event regulated by CD11b/CD18.
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PMID:Phosphorylation of complement component C3 after synthesis in U937 cells by a putative protein kinase, casein kinase 2, which is regulated by CD11b: evidence that membrane-bound proteases preferentially cleave phosphorylated C3. 937 24

The human DNA repair protein apurinic/apyrimidinic endonuclease (APE) is a dual-function protein that has important roles in both the repair of baseless sites that arise in DNA and in regulating the redox state of a number of proteins (Ref-1). Although previous attention has been focused on how the human APE/Ref-1 gene may be regulated at the DNA level, we have instead examined if APE/Ref-1 is phosphorylated, and if so how it may affect DNA repair activity. We demonstrate here that APE/Ref-1 is indeed a substrate for phosphorylation by the serine/threonine casein kinases (CK) I and II and protein kinase C. Notably, although phosphorylation by CKI and protein kinase C had no effect whatsoever on the ability of APE/Ref-1 to act at abasic sites in DNA, phosphorylation by CKII completely abolished DNA repair activity. That phosphorylation was responsible for the loss of abasic repair activity was concluded from experiments showing that inactive APE/Ref-1 could be reversed to an active DNA repair protein with phosphatase treatment. These results may help to explain the mechanism by which APE/Ref-1 switches from one unrelated function to another.
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PMID:The DNA repair activity of human redox/repair protein APE/Ref-1 is inactivated by phosphorylation. 940 49

A major pathway for K+ efflux in human reticulocytes and young RBCs is K:Cl cotransport (K:Cl-CT). The activity of K:Cl-CT is increased in pathologic RBCs containing hemoglobins S and C and may contribute to the abnormal dehydration state of these cells. Human K:Cl-CT (gene product KCC1) has been recently sequenced from human (hKCC1), rabbit and rat tissue by Gillen et al. (J Biol Chem 271:16237, 1996). We report here the sequence of KCC1 from human and mouse erythroleukemic cells (K562 and MEL cells, respectively). The cDNA for human erythroid-KCC1 is 100% identical to hKCC1 and the cDNA for mouse erythroid-KCC1 shares 89% identity with hKCC1, which translates to 96% identity at the amino acid level. Mammalian KCC1 is strongly conserved with >95% identity between human, rabbit, rat, and mouse KCC1 proteins. We did not detect any full-length mRNA transcripts of human erythroid-KCC1 in circulating reticulocytes. We detected two mRNA isoforms of human erythroid-KCC1 that resulted in C-terminal truncated proteins (73 amino acid and 17 amino acids, respectively). Human and mouse erythroidKCC1 differed at several consensus sites including a predicted PKC phosphorylation site at 108threonine and a predicted CK2 phosphorylation site at 51serine, within the predicted cytoplasmic N-terminal, that are present in human but not mouse erythroid-KCC1. Expression of MEL-KCC1 mRNA increases substantially upon DMSO-induced differentiation opening the possibility that erythroid-KCC1 plays a role in early erythroid maturation events. The molecular identification of erythroid-KCC1 is an important step towards understanding the physiologic role mediated by this protein in young and pathologic RBCs and during erythropoiesis, as well as providing a new tool for the elucidation of pathways and signals involved in RBC volume regulation.
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PMID:Molecular identification and expression of erythroid K:Cl cotransporter in human and mouse erythroleukemic cells. 951 79

A biochemical approach was used to identify proteins which interact with human BRCA1. Through this work, a kinase activity which co-purifies with BRCA1 has been identified. This kinase activity, which phosphorylates BRCA1 in vitro, was originally identified in Sf9 insect cells but is also present in cells of human origin including breast and ovarian carcinoma cell lines. The BRCA1 kinase activity in vitro is associated with a fragment of BRCA1 encompassing amino acids 329-435. This peptide is also phosphorylated in various human cell lines. A computer-assisted sequence analysis revealed that this peptide was a potential substrate for phosphorylation by PKA, PKC, or CKII. However, phosphorylation by these kinases could not be demonstrated in vitro indicating the presence of another kinase activity. Phosphorylation in vitro requires a minimal domain of BRCA1 encompassing amino acids 379-408. Notably, deletion of this minimal domain abolishes growth suppression by BRCA1 indicating that this domain, as well as phosphorylation within this domain, may be important for BRCA1 function.
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PMID:Identification of a BRCA1-associated kinase with potential biological relevance. 951 77

Calnexin is a lectin-like chaperone of the endoplasmic reticulum (ER) that couples temporally and spatially N-linked oligosaccharide modifications with the productive folding of newly synthesized glycoproteins. Calnexin was originally identified as a major type I integral membrane protein substrate of kinase(s) associated with the ER. Casein kinase II (CK2) was subsequently identified as an ER-associated kinase responsible for the in vitro phosphorylation of calnexin in microsomes (Ou, W-J., Thomas, D. Y., Bell, A. W., and Bergeron, J. J. M. (1992) J. Biol. Chem. 267, 23789-23796). We now report on the in vivo sites of calnexin phosphorylation. After 32PO4 labeling of HepG2 and Madin-Darby canine kidney cells, immunoprecipitated calnexin was phosphorylated exclusively on serine residues. Using nonradiolabeled cells, we subjected calnexin immunoprecipitates to in gel tryptic digestion followed by nanoelectrospray mass spectrometry employing selective scans specific for detection of phosphorylated fragments. Mass analyses identified three phosphorylated sites in calnexin from either HepG2 or Madin-Darby canine kidney cells. The three sites were localized to the more carboxyl-terminal half of the cytosolic domain: S534DAE (CK2 motif), S544QEE (CK2 motif), and S563PR. We conclude that CK2 is a kinase that phosphorylates calnexin in vivo as well as in microsomes in vitro. Another yet to be identified kinase (protein kinase C and/or proline-directed kinase) is directed toward the most COOH-terminal serine residue. Elucidation of the signaling cascade responsible for calnexin phosphorylation at these sites in vivo may define a novel regulatory function for calnexin in cargo folding and transport to the ER exit sites.
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PMID:Conserved in vivo phosphorylation of calnexin at casein kinase II sites as well as a protein kinase C/proline-directed kinase site. 964 93

Two distinct regions of loss of heterozygosity (LOH) in breast carcinomas were previously identified at chromosome 11q23. With the aim of identifying a tumor suppressor gene, we undertook the isolation and characterization of genes within LOH region 2, defined between loci D11S1345 and D11S1316, which spans an area of about 1 Mb. Here, we describe the cloning and characterization of a new gene, ZNF202. The gene, which spans a genomic area of approximately 10 kb, is almost exclusively expressed in testis as a 4-kb mRNA. The predicted amino acid sequence of the protein product revealed significant homologies with zinc finger proteins, indicating that the ANF202 protein may function as a transcription factor. The presence of multiple CK2 and PKC phosphorylation sites suggests that its activity may be regulated by phosphorylation. The gene is also expressed in breast carcinoma cell lines. However, mutation analysis of 39 breast cancer samples revealed no evidence of mutations, indicating that ZNF202 is unlikely to be involved in the pathogenesis of this neoplasm. Nevertheless, a role for ZNF202 in the tumorigenic process of other tissues cannot be excluded.
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PMID:Molecular cloning and characterization of ZNF202: a new gene at 11q23.3 encoding testis-specific zinc finger proteins. 979 Jul 54


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