Phosphorylation of fibrinogen by casein kinase 2

Guasch, Maria D.; Plana, Maria; Pena, J. Manuel; Itarte, Emilio

doi:10.1042/bj2340523

Cited by 18 publications

(7 citation statements)

References 25 publications

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“…The most likely and unique candidate for phosphate incorporation within this region (amino acid region 680 -709) is Ser 692 . These data are consistent with previous findings (21) and clearly demonstrate that factor Va heavy chain is a physiological substrate for platelet-mediated phosphorylation as is also the case for fibrinogen (45).…”

Section: Ckii-mediated Phosphorylation Of Human Factor V Localizationsupporting

confidence: 93%

Identification and Partial Characterization of Factor Va Heavy Chain Kinase from Human Platelets

Kalafatis

1998

Journal of Biological Chemistry

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Factor Va, the essential cofactor for prothrombinase, is phosphorylated on the acidic COOH terminus of the heavy chain of the cofactor, at Ser 692 , by a platelet membrane-associated casein kinase II (CKII). Consistent with this observation, phosphorylation of the factor Va heavy chain by the platelet kinase was inhibited by heparin. The membrane-associated platelet CKII kinase was partially purified using heparin-agarose, phosphocellulose, and ion exchange chromatography. CKII antigen was monitored using a polyclonal antibody to the ␣-subunit, and kinase activity in the various fractions was confirmed using human factor Va as a substrate. Immunoblotting experiments using polyclonal antibodies raised against synthetic peptides mimicking a portion of the deduced amino acid sequence of the ␣-, ␣ -, and ␤-subunits of human CKII demonstrated the coexistence of both ␣-and ␣ -subunits in platelets and suggested that the platelet CKII kinase may exist in part as an ␣␣ ␤ 2 complex. It is also possible that there are two distinct populations of CKII in platelets, one that is ␣␣/␤␤ and one that is ␣ ␣ /␤␤. In the presence of the purified platelet-derived CKII, human factor Va incorporates between 0.8 and 1.3 mol of phosphate/mol of factor Va depending on the concentration of the ␤-subunit in the kinase preparation. A peptide mimicking the sequence 687-705 of the human factor V molecule incorporates radioactivity in the presence of purified CKII and inhibits factor Va heavy chain phosphorylation by the platelet CKII. In contrast, a peptide with an alanine instead of a serine at position 692 neither incorporates phosphate nor inhibits factor Va phosphorylation by the platelet CKII. Human factor Va is inactivated by activated protein C following three cleavages of the heavy chain at Arg 506 , Arg 306 , and Arg 679 . Cleavage at Arg 506 is necessary for efficient exposure of the inactivating cleavage site at Arg 306 . The phosphorylated cofactor has increased susceptibility to inactivation by activated protein C, since phosphorylated factor Va was found to be inactivated approximately 3-fold faster than its native counterpart. Acceleration of the inactivation process of the phosphorylated cofactor occurs because of acceleration of the rate of cleavage at Arg 506 . These data suggest a critical role for factor Va phosphorylation on the surface of platelets in regulating cofactor activity.Coagulation factor Va is the cofactor for prothrombinase (1, 2). Factor Va combines with the serine protease factor Xa on a phospholipid or platelet surface in the presence of divalent cations to catalyze the conversion of prothrombin to thrombin. Factor Xa alone can activate prothrombin to ␣-thrombin. However, the binding of factor Va to factor Xa on a membrane surface in the presence of Ca 2ϩ increases the efficiency of the complex by 5 orders of magnitude as compared with factor Xa alone (3).Most of the factor V is found in the plasma, where it circulates at a concentration of 7 g/ml (20 nM). Approximately 20% of the total human factor V fo...

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Section: Ckii-mediated Phosphorylation Of Human Factor V Localizationsupporting

confidence: 93%

Identification and Partial Characterization of Factor Va Heavy Chain Kinase from Human Platelets

Kalafatis

1998

Journal of Biological Chemistry

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“…The remaining peptides consisted of the three FPA fragments: FPA -70%, FPA-P -20%, desAla FPA -10% (21). Previous studies with fibrinogen have demonstrated that phosphorylation of the Aa chain serine, a post-translational modification occurring prior to secretion from the liver (22)(23)(24), and removal of the amino-terminal alanine from the Aa chain are regulated with respect to subject parameters, such as age, trauma, or illness (24)(25)(26)(27), and are in place on fibrinogen prior to thrombin action (22)(23)(24). The low levels of FPA present in EDTA plasma can be attributed to the expectedly minimal thrombin activity in vivo and minimal ex vivo thrombin activation accomplished by calcium sequestration by EDTA.…”

Section: Fpa Peptides In Serummentioning

confidence: 99%

Thrombin induces broad spectrum proteolysis in human serum samples

O'Mullan¹,

Craft²,

Yi³

et al. 2009

Clinical Chemistry and Laboratory Medicine

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Background: During clotting, a thrombin cleaves fibrinogen releasing fibrinopeptide A (FPA). FPA is easily identified in serum using matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry (MS). Using MALDI-TOF MS, we observed multiple, progressively shorter fragments of serum FPA. Following ambient incubation of serum, variations in the content of FPA fragments occur over time. Denaturation of a thrombin by heating the serum sample appears to minimize this variation. These observations suggest that intrinsic proteolytic and peptidolytic activity is elevated in serum and perhaps originates from the coagulation cascade enzymes themselves, especially a thrombin. Methods: Extrinsic addition of a thrombin to a subset (3-30 kDa) of plasma proteins was carried out to induce proteolysis and to examine the resultant peptides to reveal a thrombin susceptible parent proteins. One of these identified proteins, hemopexin, was directly digested by a thrombin and the peptides examined to confirm the observations from the initial plasma protein digestion. Results: Extrinsic addition of a thrombin to a subset (3-30 kDa) of plasma proteins results in wide-spread digestion of proteins unrelated to coagulation, revealing a substrate range encompassing more than fibrinogen. Direct digestion of one of these proteins, hemopexin, by a thrombin confirms these observations. Conclusions: The resulting peptides indicate broad tolerance beyond the consensus R-G cleavage site of fibrinogen; in fact, there appears to be no bias for the amino acid following the R/K residue. These data support our hypothesis that the enzymatic activities inherent to coagulation, or at least to thrombin, contribute to destabilization of the protein and peptide content of serum. Clin Chem Lab Med 2009;47:685-93.

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“…The protein tyrosine kinase (~40) was purified from bovine thymus as described by Zioncheck et al [32]. Casein kinase-1 was obtained from rat liver cytosol as indicated in [34].…”

Section: Enzyme Purification and Assaymentioning

confidence: 99%

Phosphorylation and activation of p40 tyrosine kinase by casein kinase‐1

et al. 1990

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Because examination of regulatory rrans-phosphorylations can help elucidate the cellular functions of tyrosyl protein kinases, we have investigated the effects of phosphorylation by casein kinase-I on the activity of the p40 tyrosyl protein kinase. We find that casein kinase-1 can phosphorylate the p40 tyrosyl kinase on serine and threonine residues, in part on a unique tryptic peptide. The phosphorylation induces a substantial increase in the tyrosyl protein kinase activity of p40, in contrast to most instances in which serine/threonine phosphorylation inhibits activity of tyrosyl protein kinases. These findings raise the possibility that p40 might be part of a protein phosphorylation network in which casein kinase-I participates.

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Phosphorylation of fibrinogen by casein kinase 2

Cited by 18 publications

References 25 publications

Identification and Partial Characterization of Factor Va Heavy Chain Kinase from Human Platelets

Identification and Partial Characterization of Factor Va Heavy Chain Kinase from Human Platelets

Thrombin induces broad spectrum proteolysis in human serum samples

Phosphorylation and activation of p40 tyrosine kinase by casein kinase‐1

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