Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.1.3.1 (alkaline phosphatase)
47,916 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Antibody Ab262 was raised against a synthetic tau peptide (SKIGSTENLK, amino acids 258-267 of tau, termed Ser262 peptide). The antibody was more reactive with Ser262 peptide and unphosphorylated tau than a related phosphopeptide [SKIGS(P)TENLK, termed PSer262 peptide] and tau phosphorylated by a partially purified kinase, glycogen synthase kinase (GSK) 3 beta. AB262 reacted poorly with a peptide having the sequence DRV-QSKIGSLD (amino acids 348-358). Treatment of PSer262 peptide or GSK 3 beta phosphorylated tau with alkaline phosphatase increased Ab262 immunoreactivity, indicating that Ab262 is a reagent useful for studying tau phosphorylation at the Ser262 residue. The Ab262 immunoreactivity was detected in tau from normal brains and Alzheimer paired helical filament (PHF-tau) and in PHFs. Alkaline phosphatase treatment had no effect on the Ab262 immunoreactivity of normal tau and PHF-tau but altered the tau-1 and PHF-1 immunoreactivities, tau proteins from rat brains at 3 and 8 h postmortem exhibited 5 and 19%, respectively, more Ab262 immunoreactivity than tau from fresh tissues. In comparison, rat tau at 8 h postmortem was 40% more immunoreactive with Tau-1. The results suggest that Ser262 is not a major phosphorylation site in vivo. Moreover, there is little or no difference between PHF-tau and normal tau in the extent of phosphorylation at Ser262.
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PMID:The state of phosphorylation of normal adult brain tau, fetal tau, and tau from Alzheimer paired helical filaments at amino acid residue Ser262. 876 76

Previous studies have shown that the levels of the microtubule-associated protein tau in the CSF of patients with Alzheimer's disease (AD) are elevated compared with age-matched controls. In spite of these findings, the nature of tau in CSF has not been well documented. In the present study, tau was immunoprecipitated from CSF of patients with AD or acute stroke, as well as normal elderly controls, followed by immunoblot analysis. In all cases, CSF tau consisted primarily of a band migrating at 26-28 kDa. In AD and stroke patients, several smaller tau fragments were also detected. No intact tau was detected in any of the CSF samples examined. Further immunoprecipitation studies showed that the majority of the tau fragments contained the amino terminus of the molecule. Treatment of CSF tau with alkaline phosphatase did not alter the electrophoretic properties of the fragments. These studies clearly demonstrate that CSF tau is truncated rather than intact.
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PMID:The tau protein in human cerebrospinal fluid in Alzheimer's disease consists of proteolytically derived fragments. 897 56

Prion diseases are disorders of protein conformation that produce neurodegeneration in humans and animals. Studies of transgenic (Tg) mice indicate that a factor designated protein X is involved in the conversion of the normal cellular prion protein (PrPC) into the scrapie isoform (PrPSc); protein X appears to interact with PrPC but not with PrPSc. To search for PrPC binding proteins, we fused PrP with alkaline phosphatase (AP) to produce a soluble, secreted probe. PrP-AP was used to screen a lambdagt11 mouse brain cDNA library, and six clones were isolated. Four cDNAs are novel while two clones are fragments of Nrf2 (NF-E2 related factor 2) transcription factor and Aplp1 (amyloid precursor-like protein 1). The observation that PrP binds to a member of the APP (amyloid precursor protein) gene family is intriguing, in light of possible relevance to Alzheimer's disease. Four of the isolated clones are expressed preferentially in the mouse brain and encode a similar motif.
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PMID:Identification of candidate proteins binding to prion protein. 917 30

Oxidative stress has been implicated in the mechanism of aging and neurodegenerative disorders such as Alzheimer's disease (AD). Menadione causes oxidative stress by generating reactive oxygen species through its redox cycling and these free radicals are detoxified subsequently at the expense of intracellular thiol homeostasis. In non-neuronal cells, the cytoskeleton is a prime target of menadione-induced thiol oxidation. We used cultured human neuroblastoma MSN cells in this study to determine how tau proteins in neuronal cells are affected by menadione exposure. Menadione caused a dose-dependent thiol oxidation in these cells just like their non-neuronal counterparts. A prominent consequence of such oxidative insult in these neuronal cells was tau dephosphorylation. This dephosphorylation resulted in disappearance of phosphorylated 57-kDa tau with a concomitant emergence of 53-kDa tau whose full-length nature is indicated by its reactivity with antibodies Alz 50, Tau-1 and Tau-46. Immunochemical analyses using phosphorylation-dependent immunoprobes Tau-1 and PHF-1 with the aid of alkaline phosphatase demonstrated that 53-kDa tau was derived from dephosphorylation of 57-kDa tau. Despite its effect on thiol oxidation, menadione treatment did not lead to cytoskeletal changes reminiscent of the neurofibrillary tangles of AD. The data thus indicate that tau dephosphorylation constitutes a major feature of the menadione-induced oxidative injury in these neuronal cells.
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PMID:Menadione-induced tau dephosphorylation in cultured human neuroblastoma cells. 923 26

In Alzheimer's disease (AD) the microtubule-associated protein tau is excessively phosphorylated in degenerating neurons, but the mechanisms underlying the increased phosphorylation are unknown. Recent findings suggest that oxidative stress, and membrane lipid peroxidation in particular, contributes to the neurodegenerative process in AD. We now report that following exposure of cultured rat hippocampal neurons to 4-hydroxynonenal (HNE), an aldehydic product of membrane lipid peroxidation, tau is resistant to dephosphorylation. Immunocytochemical and Western blot analyses using phosphorylation-sensitive tau antibodies showed that HNE treatment causes a moderate increase in basal levels of tau phosphorylation, and prevents tau dephosphorylation by alkaline phosphatase in neurons pretreated with the phosphatase inhibitor okadaic acid. Studies with anti-HNE antibodies showed that HNE binds directly to tau, and that HNE immunoreactivity localizes to cell bodies and axons, cell compartments that contain tau. These data suggest a role for HNE in altered tau phosphorylation and neurofibrillary degeneration in AD.
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PMID:4-Hydroxynonenal, a product of lipid peroxidation, inhibits dephosphorylation of the microtubule-associated protein tau. 924 25

AD66 proteins derived from sodium dodecylsulfate (SDS) insoluble paired helical filaments (PHF) were isolated from Alzheimer's brain using a purification procedure developed previously in this laboratory, and characterized by immunologic and chemical cleavage methods. AD66 proteins were immunoreactive with antibodies that recognize the amino terminal, tubulin-binding, and carboxy terminal domains of microtubule-associated protein tau indicating the presence of the entire tau sequence in AD66 proteins. These proteins were reactive with antibody 423 that binds to PHF but not human adult tau. Immunologic and chemical cleavage studies indicated that only two of the six tau isoforms were present in these proteins. AD66 proteins were comprised of tau proteins containing only three tubulin binding domains with either a 29 amino acid insert or no amino terminal insert. For comparative purposes, SDS soluble PHF-tau (A68 proteins) was purified from Alzheimer's brains and normal adult tau purified from control brains. Antibody Alz-50 was immunoreactive with PHF-tau or normal tau regardless of alkaline phosphatase treatment while immunoreactivity was only observed with dephosphorylated AD66 proteins. A second phosphorylated epitope on AD66 proteins but not PHF-tau or normal tau proteins was demonstrated with antibody PHF9. These data suggest that AD66 proteins represent a more phosphorylated form of tau than PHF-tau or normal tau proteins. Two-dimensional gel electrophoresis demonstrated that AD66 proteins have higher apparent molecular weights and lower pI values than normal tau, differences possibly due to the greater phosphorylation observed in these proteins.
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PMID:Identification of microtubule-associated protein tau isoforms in Alzheimer's paired helical filaments. 925 Jun 24

The glial cytoplasmic inclusion (GCI) is a histological hallmark for multiple system atrophy (MSA). These inclusions are in oligodendrocytes, contain microtubular structures of 20-30 nm diameter, and can be labelled immunohistochemically with antibodies to ubiquitin, alphaB-crystallin, alpha- and beta-tubulin, and the microtubule-associated protein tau. GCIs have been compared with neuronal inclusions in other neurodegenerative disorders including the neurofibrillary tangles (NFTs) found in Alzheimer's disease (AD), which also contain tau protein. In order to determine whether the tau protein of GCIs in MSA is similar to that observed in AD we used a panel of antibodies to phosphorylation-independent (SMI51, TP007, TP70), dephosphorylation-dependent (Tau.1), and phosphorylation-dependent antibodies to tau and neurofilaments (AT8, AT180, AT270, SMI31, SMI34, RT97, BF10, 8D8). Immunohistochemistry was performed on paraffin wax-embedded brain tissue of the cerebellum, brainstem, and frontal lobes (Brodmann areas 4/6) of ten clinically and neuropathologically well-characterised cases of MSA, two cases of AD, and two normal controls. The NFTs of the AD cases were labelled with all the phosphorylation-dependent and phosphorylation-independent antibodies and with Tau.1 only after treatment with alkaline phosphatase. In contrast, GCIs were immunolabelled by the phosphorylation-independent antibodies and Tau.1, but not by the phosphorylation-dependent antibodies. These data demonstrate that the tau in GCIs is different from the abnormally phosphorylated tau found in AD and is similar to normal adult tau. The mechanism causing the abnormal accumulation of tau in GCIs remains to be elucidated.
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PMID:Tau protein in the glial cytoplasmic inclusions of multiple system atrophy can be distinguished from abnormal tau in Alzheimer's disease. 925 61

Microtubules are important for the growth and maintenance of stable neuronal processes and their organisation is controlled partly by microtubule-associated proteins (MAPs). MAP 1B is the first MAP to be expressed in neurons and plays an important role in neurite outgrowth. MAP 1B is phosphorylated at multiple sites and it is believed that the function of the protein is regulated by its phosphorylation state. We have shown that the monoclonal antibody (mAb) RT97, which recognises phosphorylated epitopes on neurofilament proteins, fetal tau, and on Alzheimer's paired helical filament-tau, also recognises a developmentally regulated phosphorylation epitope on MAP 1B. In the rat cerebellum, Western blot analysis shows that mAb RT97 recognises the upper band of the MAP 1B doublet and that the amount of this epitope peaks very early postnatally and decreases with increasing age so that it is absent in the adult, despite the continued expression of MAP 1B in the adult. We confirmed that mAb RT97 binds to MAP 1B by showing that it recognises MAP 1B immunoprecipitated from postnatal rat cerebellum using polyclonal antibodies to recombinant MAP 1B proteins. We established that the RT97 epitope on MAP 1B is phosphorylated by showing that antibody binding was abolished by alkaline phosphatase treatment of immunoblots. Epitope mapping experiments suggest that the mAb RT97 site on MAP 1B is near the N-terminus of the molecule. Despite our immunoblotting data, immunostaining of sections of postnatal rat cerebellum with mAb RT97 shows a staining pattern typical of neurofilaments with no apparent staining of MAP 1B. For instance, basket cell axons and axons in the granule cell layer and white matter stained, whereas parallel fibres did not. These results suggest that the MAP 1B epitope is masked or lost under the immunocytochemical conditions in which the cerebellar sections are prepared. The upper band of the MAP 1B doublet is believed to be predominantly phosphorylated by proline-directed protein kinases (PDPKs). PDPKs are also good candidates for phosphorylating neurofilament proteins and tau and therefore we postulate that the sites recognised by RT97 on these neuronal cytoskeletal proteins may be phosphorylated by similar kinases. Important goals are to determine the precise location of the RT97 epitope on MAP 1B and the kinase responsible.
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PMID:The neurofilament antibody RT97 recognises a developmentally regulated phosphorylation epitope on microtubule-associated protein 1B. 930 99

The chemical interaction that condenses the hyperphosphorylated protein tau in Alzheimer's disease (AD P-tau) into neurofibrillary tangles and cripples synaptic transmission remains unknown. Only beta-sheet, positive ion salt bridges between phosphates, and hydrophobic association can create tangles of just AD P-tau. We have correlated transmission electron microscope (TEM) images of tau aggregation with different percentages of beta-sheet in aqueous suspensions of tau while using buffers that block dispositive or tripositive ionic bridges between intermolecular phosphates. Circular dichroism (CD) studies were performed at different temperatures from 5-85 degrees C using AD P-tau, AD P-tau dephosphorylated with hydrofluoric acid (HF AD P-tau) or alkaline phosphatase (AP AD P-tau), and recombinant human tau with 3-repeats and two amino terminal inserts (R-39) and using bovine tau (B tau) isolated without heat or acid treatment. Secondary structure was estimated from CD spectra at 5 degrees C using the Lincomb algorithm. Each preparation except one demonstrated an inverse temperature transition, Ti, in the CD at 197 nm. No correlation was found between beta-sheet content and aggregation, leaving only hydrophobic interaction as the remaining possibility. Thirteen of 21 possible phosphorylation sites in AD P-tau lie adjacent to positive residues in tau's primary structure. Occupation of five to nine phosphate sites on AD P-tau appears sufficient to reduce or neutralize tau's basic character. AD P-tau's hydrophobic character is indicated by its low inverse temperature transition, Ti. The Ti for AD P-tau was 24.5 degrees C or 28 degrees C, whereas for B tau with three phosphates it was 32 degrees C, for unphosphorylated tau R-39 it was 38 degrees C, and for dephosphorylated HF AD P-tau it was 37.5 degrees C. The hydrophobic protein elastin and its analogs coalesce and precipitate at their Ti of 24-29 degrees C, well below body temperature. We hypothesize that AD P-tau causes tangle accumulation by this mechanism.
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PMID:Alzheimer disease hyperphosphorylated tau aggregates hydrophobically. 932 57

We have been using alkaline phosphatase (AP) histochemical staining, formerly a research tool for the study of cerebral cortical vascular morphology, to examine pathological changes in the cortex and deep cerebral structures. Deep structures stain similarly to the cortex. The AP stain is found in the afferent vessels (small arteries, arterioles, and capillaries), but not in venules and veins. The stain is also present in leaky vessels, such as those in the area postrema. The vascular supply to the cerebrum is not homogeneous. Supply to the deep white matter, for instance, derives from the leptomeningeal border zone, and then medullary arterioles must wind their way for up to 4 cm before arriving at their ultimate destination. Adding to the difficulties, tortuosities develop in some of these vessels with aging. According to some calculations, hypertensive levels of blood pressure would be required to maintain irrigation through some of these vessels. We have identified a venous alteration that attends aging: periventricular venous collagenosis (PVC) is a previously unrecognized, noninflammatory, mural disease of the periventricular veins. In severe cases, examples can be found of veins that are completely occluded by this process. PVC is found in 65% of subjects over 60 years old, and it strongly correlates with leukoaraiosis. In addition to previously mentioned aging-related changes, we have found extreme tortuosity, multiplications, and aneurysms of the smallest arterioles and lumpy-bumpy capillaries in the deep structures of patients with Alzheimer's disease.
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PMID:Cerebral microvascular alterations in aging, leukoaraiosis, and Alzheimer's disease. 932 84


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