共查询到20条相似文献,搜索用时 15 毫秒
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Thaís Posser† Jeferson L. Franco† Larisa Bobrovskaya Rodrigo B. Leal† Phillip W. Dickson Peter R. Dunkley 《Journal of neurochemistry》2009,110(3):848-856
Manganese (Mn2+ ) is an essential metal involved in normal functioning of a range of physiological processes. However, occupational overexposure to Mn2+ causes neurotoxicity. The dopaminergic system is a particular target for Mn2+ neurotoxicity. Tyrosine hydroxylase (TH) is the rate limiting enzyme for dopamine synthesis and is regulated acutely by phosphorylation at Ser40 and chronically by protein synthesis. In this study we used pheochromocytoma 12 cells to investigate the effects of Mn2+ exposure on the phosphorylation and activity of TH. Mn2+ treatment for 24 h caused a sustained increase in Ser40 phosphorylation and TH activity at a concentration of 100 μM, without altering the level of TH protein or PC12 cell viability. Inhibition of protein kinase A and protein kinase C and protein kinases known to be involved in sustained phosphorylation of TH in response to other stimuli did not block the effects of Mn2+ on Ser40 phosphorylation. A substantial increase in H2 O2 production occurred in response to 100 μM Mn2+ . The antioxidant TroloxTM completely inhibited H2 O2 production but did not block TH phosphorylation at Ser40, indicating that oxidative stress was not involved. Sustained TH phosphorylation at Ser40 and the consequent activation of TH both occurred at low concentrations of Mn2+ and this provides a potential new mechanism for Mn2+ -induced neuronal action that does not involve H2 O2 -mediated cell death. 相似文献
3.
Abstract: The cellular localization of two Ca2+ -dependent protein phosphorylation systems was investigated using the kainic acid lesioning technique for the selective destruction of neurons. In one of these systems, a crude synaptosomal (P2 ) fraction was preincubated with 32 Pj for 30 min; the phosphorylation of several proteins was increased during a short subsequent incubation with veratridine plus Ca2+ . In the second system, crude synaptosomal membranes isolated from the P2 fraction were incubated with [γ-32 P]ATP; in this system, the phosphorylation of several proteins was increased in the presence of a "calcium-dependent regulator" plus Ca2+ . Kainic acid lesioning greatly reduced the amount of Ca-+ -dependent protein phosphorylation in both systems. The results indicate a predominantly neuronal localization for both Ca2+ -dependent protein phosphorylation systems. 相似文献
4.
Steven R. Ennis Nobuyuki Kawai Xiao-dan Ren Galaleldin E. Abdelkarim Richard F. Keep 《Journal of neurochemistry》1998,71(6):2565-2573
Abstract: The aim of this study was to determine the effect of angiotensin II (AII) on tyrosine hydroxylase (TOH) activity and phosphorylation in bovine adrenal chromaffin cells (BACCs). We report here that stimulation of BACCs with AII (100 n M ) produced a significant increase in both TOH activity and phosphorylation over a period of 10 min. The increase in TOH activity was receptor-mediated. Tryptic phosphopeptide analysis by HPLC revealed that AII stimulated an increase in phosphorylation of three sites on TOH, Ser19 , Ser31 , and Ser40 , with the largest increase being observed for Ser31 phosphorylation. Pretreatment of the cells with the protein kinase C inhibitor Ro 31-8220 (10 µ M , 15 min) did not affect TOH activity or phosphorylation produced by AII. The inhibitor also did not affect the TOH activity or Ser40 phosphorylation produced by forskolin (10 µ M , 10 min). In contrast, Ro 31-8220 fully inhibited the TOH activation as well as Ser31 and Ser40 phosphorylation of TOH produced by phorbol 12, 13-dibutyrate (500 n M , 10 min). Removal of extracellular Ca2+ from the incubation medium inhibited the AII-induced TOH activity by 50% and significantly blocked Ser19 and Ser31 phosphorylation but did not affect Ser40 phosphorylation in response to AII. These results indicate that AII activates a complex and perhaps novel signaling pathway leading to the phosphorylation and activation of TOH. The TOH activation by AII appears to be partially independent of Ser40 phosphorylation, suggesting a potentially important role for Ser31 phosphorylation. 相似文献
5.
F. M. J. Heemskerk L. H. Schrama W. E. J. M. Ghijsen P. N. E. De Graan F. H. Lopes da Silva W. H. Gispen 《Journal of neurochemistry》1991,56(6):1827-1835
Abstract: Recently we have shown that 4-aminopyridine (4-AP), a drug known to enhance transmitter release, stimulates the phosphorylation of the protein kinase C substrate B-50 (GAP-43) in rat brain synaptosomes and that this effect is dependent on the presence of extracellular Ca2+ . Hence, we were interested in the relationship between changes induced by 4-AP in the intracellular free Ca2+ concentration ([Ca2+ ]i ) and B-50 phosphorylation in synaptosomes. 4-AP (100 μ M ) elevates the [Ca2+ ]i (as determined with fura-2) to approximately the same extent as depolarization with 30 m M K+ (from an initial resting level of 240 n M to ∼480 n M after treatment). However, the underlying mechanisms appear to be different: In the presence of 4-AP, depolarization with K+ still evoked an increase in [Ca2+ ]i , which was additive to the elevation caused by 4-AP. Several Ca2+ channel antagonists (CdCl2 , LaCl3 , and diphenylhydantoin) inhibited the increase in B-50 phosphorylation by 4-AP. It is interesting that the increase in [Ca2+ ]i and the increase in B-50 phosphorylation by 4-AP were attenuated by tetrodotoxin, a finding pointing to a possible involvement of Na+ channels in this action. These results suggest that 4-AP (indirectly) stimulates both Ca2+ influx and B-50 phosphorylation through voltage-dependent channels by a mechanism dependent on Na+ channel activity. 相似文献
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Abstract The Escherichia coli ptsI and ptsH genes code for the synthesis of two proteins of the phosphoenolpyruvate-dependent phosphotransferase system (PTS), namely enzyme I and protein HPr. A number of ptsI + ptsH + /F' ptsI + ptsH merodiploids was obtained. It was shown in experiments in vivo that ptsH mutations in the transposition are dominant. Bacterial extracts from these merodiploids supported [14 C]methyl glucoside (MG) phosphorylation at the expense of phosphoenolpyruvate only half as much as extracts from the pts + cells. ptsI + ptsH /F' ptsI + ptsH + merodiploids appeared to be non-viable; the reason for this lack of viability is discussed. 相似文献
7.
Else Marie Fykse 《Journal of neurochemistry》1998,71(4):1661-1669
Abstract: Studies performed over the past several years have provided evidence that phosphorylation of proteins is important in the regulation of neurotransmitter release. In this study, it is shown that rabphilin-3A is present in cerebellar granule cells as a phosphoprotein, by using 32 P-labeling of cerebellar granule cells, immunoprecipitation, phosphoamino acid analysis, and phosphopeptide mapping. The level of phosphorylation was increased (224 ± 13%) (mean ± SEM) on depolarization of the cells with K+ (56 m M ) in the presence of external Ca2+ (1 m M ). Stimulation of protein kinase C with a phorbol ester (phorbol 12,13-dibutyrate) also enhanced the phosphorylation of rabphilin-3A (217 ± 21%). Inhibitors of Ca2+ /calmodulin-stimulated protein kinases or protein kinase C reduced the depolarization-enhanced phosphorylation of rabphilin-3A, indicating that rabphilin-3A is one of the targets for Ca2+ -activated protein kinases in the nerve terminal. Costimulation of cells with phorbol 12,13-dibutyrate and K+ depolarization produced an increased level of phosphorylation of rabphilin-3A compared with either stimulus alone (287 ± 61%). Phosphoamino acid analysis showed that serine was the main phosphorylated residue. A slight increase in the threonine phosphorylation could also be detected, whereas tyrosine phosphorylation could not be detected at all. These results suggest that rabphilin-3A is phosphorylated in vivo and undergoes synaptic activity-dependent phosphorylation during Ca2+ -activated K+ depolarization. 相似文献
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Carlos-Alberto Gonçalves Amanda Hall Alistair T. R. Sim Stephen J. Bunn Philip D. Marley Tat B. Cheah Peter R. Dunkley 《Journal of neurochemistry》1997,69(6):2387-2396
Abstract: The protein kinases and protein phosphatases that act on tyrosine hydroxylase in vivo have not been established. Bovine adrenal chromaffin cells were permeabilized with digitonin and incubated with [γ-32 P]ATP, in the presence or absence of 10 µ M Ca2+ , 1 µ M cyclic AMP, 1 µ M phorbol dibutyrate, or various kinase or phosphatase inhibitors. Ca2+ increased the phosphorylation of Ser19 and Ser40 . Cyclic AMP, and phorbol dibutyrate in the presence of Ca2+ , increased the phosphorylation of only Ser40 . Ser31 and Ser8 were not phosphorylated. The Ca2+ -stimulated phosphorylation of Ser19 was incompletely reduced by inhibitors of calcium/calmodulin-stimulated protein kinase II (46% with KN93 and 68% with CaM-PKII 273–302), suggesting that another protein kinase(s) was contributing to the phosphorylation of this site. The Ca2+ -stimulated phosphorylation of Ser40 was reduced by specific inhibitors of protein kinase A (56% with H89 and 38% with PKAi 5–22 amide) and protein kinase C (70% with Ro 31-8220 and 54% with PKCi 19–31), suggesting that protein kinases A and C contributed to most of the phosphorylation of this site. Results with okadaic acid and microcystin suggested that Ser19 and Ser40 were dephosphorylated by PP2A. 相似文献
9.
Souad Hilmi Michel Fournier Henri Valeins Jean Claude Gandar Jacques Bonnet 《Journal of neurochemistry》1995,64(2):902-907
Abstract: Myelin membrane prepared from mouse sciatic nerve possesses both kinase and substrates to incorporate [32 P]PO4 3− from [γ-32 P]ATP into protein constituents. Among these, P0 glycoprotein is the major phosphorylated species. To identify the phosphorylated sites, P0 protein was in vitro phosphorylated, purified, and cleaved by CNBr. Two 32 P-phosphopeptides were isolated by HPLC. The exact localization of the sequences around the phosphorylated sites was determined. The comparison with rat P0 sequence revealed, besides a Lys172 to Arg substitution, that in the first peptide, two serine residues (Ser176 and Ser181 ) were phosphorylated, Ser176 appearing to be modified subsequently to Ser181 . In the second peptide, Ser197 , Ser199 , and Ser204 were phosphorylated. All these serines are clustered in the C-terminal region of P0 protein. This in vitro study served as the basis for the identification of the in vivo phosphorylation sites of the C terminal region of P0 . We found that, in vivo, Ser181 and Ser176 are not phosphorylated, whereas Ser197 , Ser199 , Ser204 , Ser208 , and Ser214 are modified to various extents. Our results strongly suggest that the phosphorylation of these serine residues alters the secondary structure of this domain. Such a structural perturbation could play an important role in myelin compaction at the dense line level. 相似文献
10.
Gregory A. Jicha Audrey O' Donnell† Charles Weaver‡ Ruth Angeletti† & Peter Davies‡ 《Journal of neurochemistry》1999,72(1):214-224
Abstract : Immunoaffinity-purified paired helical filaments (PHFs) from Alzheimer's disease (AD) brain homogenates contain an associated protein kinase activity that is able to induce the phosphorylation of PHF proteins on addition of exogenous MgCl2 and ATP. PHF kinase activity is shown to be present in immunoaffinity-purified PHFs from both sporadic and familial AD, Down's syndrome, and Pick's disease but not from normal brain homogenates. Although initial studies failed to show that the kinase was able to induce the phosphorylation of tau, additional studies presented in this article show that only cyclic AMP-dependent protein kinase-pretreated recombinant tau is a substrate for the PHF kinase activity. Deletional mutagenesis, phosphopeptide mapping, and site-directed mutagenesis have identified the PHF kinase phosphorylation sites as amino acids Thr361 and Ser412 in htau40. In addition, the cyclic AMP-dependent protein kinase phosphorylation sites that direct the PHF kinase have been mapped to amino acids Ser356 and Ser409 in htau40. Additional data demonstrate that these hierarchical phosphorylations in the extreme C terminus of tau allow for the incorporation of recombinant tau into exogenously added AD-derived PHFs, providing evidence that certain unique phosphorylations of tau may play a role in the pathogenesis of neurofibrillary pathology in AD. 相似文献
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Abstract: In this report, the phosphorylation sites of neurofilament protein of medium molecular mass (NF-M) by protein kinase FA /glycogen synthase kinase 3α (kinase FA /GSK-3α) were determined by two-dimensional electrophoresis/TLC, phosphoamino acid analysis, HPLC, Edman degradation, and peptide sequencing. Kinase FA /GSK-3α phosphorylates NF-M predominantly on serine, residue. Three major tryptic phosphopeptide peaks were resolved by C18 reverse-phase HPLC. Edman degradation and peptide sequence analysis revealed that AKS(p)PVSK is the phosphorylation site sequence for the first major peak. When mapping with the amino acid sequence of neurofilament, we finally demonstrate Ser603 -Pro, one of the in vivo sites in NF-M, as the major site phosphorylated by kinase FA /GSK-3α. By using the same approach, we also identified the in vivo sites of Ser502 -Pro, Ser506 -Pro, and Ser666 -Pro as the other three major sites in NF-M phosphorylated by kinase FA /GSK-3α. Taken together, the results provide initial evidence that kinase FA /GSK-3α may represent a physiologically relevant protein kinase involved in the in vivo phosphorylation of NF-M. Because Ser502 , Ser506 , Ser603 , and Ser666 are all flanked by a carboxyl-terminal proline residue, the results provide further evidence that FA /GSK-3α may represent a proline-directed protein kinase involved in the structure-function regulation of the neuronal cytoskeletal system. 相似文献
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Manabu Arioka Masamitsu Tsukamoto Koichi Ishiguro Rika Kato Kazuki Sato Kazutomo Imahori Tsuneko Uchida 《Journal of neurochemistry》1993,60(2):461-468
Abstract: To study the phosphorylation state of τ in vivo, we have prepared antisera by immunizing rabbits with synthetic phosphopeptides containing phosphoamino acids at specific sites that are potential targets for τ protein kinase II. Immunoblot experiments using these antisera demonstrated that τ in microtubule-associated proteins is phosphorylated at Ser144 and at Ser315 . Almost all τ variants separated on two-dimensional gel electrophoresis were phosphorylated at Ser144 and nearly one-half of them at Ser315 . Phosphorylation at Ser144 and at Thr147 of τ isolated from heat-stable brain extracts was shown to be developmentally regulated, with the highest level of phosphorylation found at postnatal week 1. In vitro phosphorylation of τ by τ protein kinase I, a kinase responsible for abnormal phosphorylation of τ found in paired helical filaments of patients with Alzheimer's disease, was enhanced by prior phosphorylation of τ by τ protein kinase II. Thus, we suggest that τ protein kinase II is indirectly involved, at least in part, in the regulation of the phosphorylation state of τ in neuronal cells. 相似文献
13.
Abstract: Soluble and membrane fractions of bovine adrenal medulla contain several substrates for the Ca2+ / phospholipid-dependent and cyclic AMP-dependent protein kinases. The phosphorylation of soluble proteins (36 and 17.7 kilodaltons) and a membrane protein (22.5 kilo-daltons) showed an absolute requirement for the presence of both Ca2+ and phosphatidylserine; other substrates showed less stringent phosphorylation requirements and many of these proteins were specific for each of the protein kinases. The Ca2+ /phospholipid-dependent phosphorylation was rapid, with effects seen as early as at 30 s of incubation. Measurement of enzyme activities with histone HI as an exogenous substrate demonstrated that the Ca2+ /phospholipid-dependent protein kinase was equally distributed between the soluble and membrane fractions whereas the cyclic AMP-dependent enzyme was predominantly membrane-bound in adrenal medulla and chromaffin cells. The activity of the soluble Ca2+ /phos-pholipid-dependent protein kinase of adrenal medulla was found to be about 50% of the enzyme level present in rat brain, a tissue previously shown to contain a very high enzyme activity. These results suggest a prominent role for the Ca2+ /phospholipid-dependent protein kinase in chromaffin cell function. 相似文献
14.
ROBERT E. SCOTT THOMAS P. DOUSA 《Differentiation; research in biological diversity》1980,16(1-3):135-140
Differences in the cyclic AMP-dependent plasma membrane phosphorylation system of undifferentiated and differentiated L6 myogenic cells have been detected. Endogenous plasma membrane protein phosphorylation in undifferentiated L6 myoblasts was stimulated more than three fold by 5 × 10−5 M cyclic AMP, whereas no statistically significant cyclic AMP-dependent phosphorylation of endogenous plasma membrane proteins was observed in differentiated L6 cells. In undifferentiated cells cyclic AMP promoted the phosphorylation of several proteins, the most prominent of which had a molecular weight of 110,000. In differentiated cells cyclic AMP did not selectively promote the phosphorylation of specific plasma membrane proteins. Both differentiated and undifferentiated L6 cells, however, contain a cyclic AMP-dependent protein kinase capable of catalyzing the phosphorylation of exogenous substrates, such as histone f2 b. Therefore, the data show that differentiation in L6 cells is associated with a selective change in the activity of a plasma membrane cyclic AMP-dependent protein kinase which employs endogenous membrane proteins as substrate. 相似文献
15.
Gerry Shaw Rehae Miller Deng-Shun Wang Dali Tang Brian A. Hollander † Gudrun S. Bennett 《Journal of neurochemistry》1997,69(4):1729-1737
Abstract: In previous studies we have identified Ser502 , Ser528 , and Ser534 as target sites in chicken neurofilament middle molecular mass protein (NF-M) for casein kinase I (CKI) in vitro and have shown that these sites are also phosphorylated in vivo. We now make use of a combination of molecular biological and protein chemical techniques to show that two additional in vivo phosphorylation sites in chicken NF-M, Ser464 and Ser471 , can also be phosphorylated by CKI in vitro. These two sites are conserved in higher vertebrate NF-M molecules, and recombinant protein constructs containing the homologous rat NF-M peptides can be phosphorylated by CKI in vitro, suggesting that phosphorylation of these sites is conserved at least in higher vertebrates. The two new sites are adjacent to a conserved peptide sequence (VEE-IIEET-V) found once in higher vertebrate NF-M molecules and twice in lamprey NF-180. Variants of this sequence are also found in neurofilament low and high molecular mass proteins (NF-L and NF-H) and α-internexin, and in mammalian NF-L are known to be associated with in vivo phosphorylation sites. We speculate that CKI phosphorylation in general, and these sites in particular, may be important in neurofilament function. 相似文献
16.
Abstract— Endogenous protein phosphorylation has been studied during in vitro polymerization of microtubules by incubating a purified tubulin preparation at 37°C in the presence of radioactive ATP. At optimal conditions the rate of phosphorylation was found to follow the course of polymerization by a shift to a lower rate at the polymerization plateau.
Zn2+ at 0.5 m m was shown to stimulate phosphorylation, mainly of tubulin-associated proteins (mol wt 110,000 and 175,000,) and to a lesser extent of tubulin. The effect occurred at Zn2+ -concentrations which induce formation of tubulin sheet polymers, which suggests that the state of aggregation of tubulin is of importance for the phosphorylation. In contrast to Zn2+ , Mg2+ only increased phosphorylation of the high molecular weight proteins, and to a lesser degree. The stimulation by Zn2+ or Mg2+ was potentiated by cyclic AMP or cyclic GMP.
A low concentration of Zn2+ (5 μ m ) or cyclic GMP at 10 μ m inhibited phosphorylation, possibly by interaction with a co-existing protein phosphatase. 相似文献
Zn
A low concentration of Zn
17.
Stephen J. Bunn Alistair T. R. Sim Lynn M. Herd Leone M. Austin Peter R. Dunkley 《Journal of neurochemistry》1995,64(3):1370-1378
Abstract: Tyrosine hydroxylase (TOH), the rate-limiting enzyme in catecholamine biosynthesis, is regulated by phosphorylation. Activation of histaminergic H1 receptors on cultured bovine adrenal chromaffin cells stimulated a rapid increase in TOH phosphorylation (within 5 s) that was sustained for at least 5 min. The initial increase in TOH phosphorylation (up to 1 min) was essentially unchanged by the removal of extracellular Ca2+ . In contrast, the H1 -mediated response was abolished by preloading the cells with BAPTA acetoxymethyl ester (50 µ M ) and significantly reduced by prior exposure to caffeine (10 m M for 10 min) to deplete intracellular Ca2+ . Trypticphosphopeptide analysis by HPLC revealed that the H1 response in the presence or absence of extracellular Ca2+ resulted in a major increase in the phosphorylation of Ser19 with smaller increases in that of Ser40 and Ser31 . In contrast, although a brief stimulation with nicotine (30 µ M for 60 s) also resulted in a major increase in Ser19 phosphorylation, this response was abolished in the absence of extracellular Ca2+ . These data indicate that the mobilization of intracellular Ca2+ plays a crucial role in supporting H1 -mediated TOH phosphorylation and may thus have a potentially important role in regulating catecholamine synthesis. 相似文献
18.
Abstract: The growth cone is responsible for axonal elongation and pathfinding by responding to various modulators for neurite growth, including neurotransmitters, although the sensor mechanisms are not fully understood. Among neurotransmitters, GABA is most likely to demonstrate activity in vivo because GABA and the GABAA receptor appear even in early stages of CNS development. We investigated the GABAA receptor-mediated signaling pathway in the growth cone using isolated growth cones (IGCs). Both the GABAA binding site and the benzodiazepine modulatory site were enriched in the growth cone membrane. In the intact IGC, GABA induced picrotoxin-sensitive Cl− flux (not influx but efflux) and increased the intracellular Ca2+ concentration in a picrotoxin- and verapamil-sensitive manner. Protein kinase C (PKC)-dependent phosphorylation of two proteins identified as GAP-43 and MARCKS protein was enhanced in the intact IGC stimulated by GABA, resulting in the release of MARCKS protein and GAP-43 from the membrane. Collectively, our results suggest the following scheme: activation of the functional GABAA receptor localized in the growth cone membrane → Cl− efflux induction through the GABAA -associated Cl− channel → Ca2+ influx through an L-type voltage-sensitive Ca2+ channel → Ca2+ -dependent phosphorylation of GAP-43 and MARCKS protein by PKC. 相似文献
19.
Rajeevkumar Raveendran Sudarsana Devi Suma Priya Madhavan Mayadevi Mathew Steephan Thankayyan R. Santhoshkumar John Cheriyan Rajendran Sanalkumar Kurup K. Pradeep Jackson James Ramakrishnapillai V. Omkumar 《Journal of neurochemistry》2009,110(1):92-105
Ca2+ influx through NMDA-type glutamate receptor at excitatory synapses causes activation of post-synaptic Ca2+ /calmodulin-dependent protein kinase type II (CaMKII) and its translocation to the NR2B subunit of NMDA receptor. The major binding site for CaMKII on NR2B undergoes phosphorylation at Ser1303, in vivo . Even though some regulatory effects of this phosphorylation are known, the mode of dephosphorylation of NR2B-Ser1303 is still unclear. We show that phosphorylation status at Ser1303 enables NR2B to distinguish between the Ca2+ /calmodulin activated form and the autonomously active Thr286-autophosphorylated form of CaMKII. Green fluorescent protein–α-CaMKII co-expressed with NR2B sequence in human embryonic kidney 293 cells was used to study intracellular binding between the two proteins. Binding in vitro was studied by glutathione- S -transferase pull-down assay. Thr286-autophosphorylated α-CaMKII or the autophosphorylation mimicking mutant, T286D-α-CaMKII, binds NR2B sequence independent of Ca2+ /calmodulin unlike native wild-type α-CaMKII. We show enhancement of this binding by Ca2+ /calmodulin. Phosphorylation or a phosphorylation mimicking mutation on NR2B (NR2B-S1303D) abolishes the Ca2+ /calmodulin-independent binding whereas it allows the Ca2+ /calmodulin-dependent binding of α-CaMKII in vitro . Similarly, the autonomously active mutants, T286D-α-CaMKII and F293E/N294D-α-CaMKII, exhibited Ca2+ -independent binding to non-phosphorylatable mutant of NR2B under intracellular conditions. We also show for the first time that phosphatases in the brain such as protein phosphatase 1 and protein phosphatase 2A dephosphorylate phospho-Ser1303 on NR2B. 相似文献
20.
Shiaw-Der Yang Jen-Shin Song Jau-Song Yu Shine-Gwo Shiah 《Journal of neurochemistry》1993,61(5):1742-1747
Abstract— Previously, we identified protein kinase FA /gly-cogen synthase kinase-3 (GSK-3) as a microtubule-associated protein kinase that can incorporate 4 mol of phosphates into 1 mol of protein and cause its electrophoretic mobility shift in sodium dodecyl sulfate gels, a unique property characteristic of paired helical filament-associated pathological (PHF-) in Alzheimer's disease brains. In this report, we identified TPPKS(p)PSAAK and SPVVSGDTS(p)PR as two phosphorylation site sequences phosphorylated by kinase FA /GSK-3 in using peptide sequence analysis and sequential manual Edman degradation for radiosequencing. When mapping with human brain sequence, we further identified Ser235 -Pro and Ser404 -Pro as the two major phosphorylation sites according to the numbering of the longest isoform. Ser235 and Ser404 have been reported as two of the major abnormal phosphorylation sites in PHF-. Taken together, the results provide initial evidence that protein kinase FA /GSK-3 may represent one of the Ser-Pro motif-directed kinases involved in the abnormal phosphorylation of pathological PHF- in Alzheimer's disease brain. 相似文献