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Human Sphingolipid Metabolic Process

Also showing Human Sphingolipid Biosynthetic Process

$260
100 µl
APPLICATIONS
REACTIVITY
Human

Application Methods: Immunoprecipitation, Western Blotting

Background: Sphingosine kinases (SPHKs) catalyze the phosphorylation of sphingosine to form sphingosine-1-phosphate (S1P), a lipid mediator with both intra- and extracellular functions. Together with other sphingolipid metabolizing enzymes, SPHKs regulate the balance of the lipid mediators, ceramide, sphingosine, and S1P (1-4). Two distinct SPHK isoforms, SPHK1 and SPHK2, have been cloned and characterized (5,6). SPHK1 and SPHK2 are highly conserved and diversely expressed (7,8). The SPHKs are activated by G protein-coupled receptors, receptor tyrosine kinases, immunoglobulin receptors, cytokines, and other stimuli (9-12). The molecular mechanisms by which SPHK1 and SPHK2 are specifically regulated are complex and only partially understood.

$260
100 µl
APPLICATIONS
REACTIVITY
Human

Application Methods: Immunoprecipitation, Western Blotting

Background: Sphingosine kinases (SPHKs) catalyze the phosphorylation of sphingosine to form sphingosine-1-phosphate (S1P), a lipid mediator with both intra- and extracellular functions. Together with other sphingolipid metabolizing enzymes, SPHKs regulate the balance of the lipid mediators, ceramide, sphingosine, and S1P (1-4). Two distinct SPHK isoforms, SPHK1 and SPHK2, have been cloned and characterized (5,6). SPHK1 and SPHK2 are highly conserved and diversely expressed (7,8). The SPHKs are activated by G protein-coupled receptors, receptor tyrosine kinases, immunoglobulin receptors, cytokines, and other stimuli (9-12). The molecular mechanisms by which SPHK1 and SPHK2 are specifically regulated are complex and only partially understood.

$122
20 µl
$303
100 µl
$717
300 µl
APPLICATIONS
REACTIVITY
Human, Monkey, Mouse, Rat

Application Methods: Immunoprecipitation, Western Blotting

Background: Activation of PKC is one of the earliest events in a cascade leading to a variety of cellular responses such as secretion, gene expression, proliferation and muscle contraction (1,2). Protein kinase D (PKD), also called PKCμ, is a serine/threonine kinase whose activation is dependent on the phosphorylation of two activation loop sites, Ser744 and Ser748, via a PKC-dependent signaling pathway (3-5). In addition to the two activation loop sites, the carboxy-terminal Ser916 has been identified as an autophosphorylation site for PKD/PKCμ. Phosphorylation at Ser916 correlates with PKD/PKCμ catalytic activity (6).

$122
20 µl
$303
100 µl
$717
300 µl
APPLICATIONS
REACTIVITY
Human, Monkey, Mouse, Rat

Application Methods: Western Blotting

Background: Activation of PKC is one of the earliest events in a cascade leading to a variety of cellular responses such as secretion, gene expression, proliferation and muscle contraction (1,2). Protein kinase D (PKD), also called PKCμ, is a serine/threonine kinase whose activation is dependent on the phosphorylation of two activation loop sites, Ser744 and Ser748, via a PKC-dependent signaling pathway (3-5). In addition to the two activation loop sites, the carboxy-terminal Ser916 has been identified as an autophosphorylation site for PKD/PKCμ. Phosphorylation at Ser916 correlates with PKD/PKCμ catalytic activity (6).

$260
100 µl
APPLICATIONS
REACTIVITY
Human, Monkey

Application Methods: Immunoprecipitation, Western Blotting

Background: Activation of PKC is one of the earliest events in a cascade leading to a variety of cellular responses such as secretion, gene expression, proliferation and muscle contraction (1,2). Protein kinase D (PKD), also called PKCμ, is a serine/threonine kinase whose activation is dependent on the phosphorylation of two activation loop sites, Ser744 and Ser748, via a PKC-dependent signaling pathway (3-5). In addition to the two activation loop sites, the carboxy-terminal Ser916 has been identified as an autophosphorylation site for PKD/PKCμ. Phosphorylation at Ser916 correlates with PKD/PKCμ catalytic activity (6).

$260
100 µl
APPLICATIONS
REACTIVITY
Human, Mouse, Rat

Application Methods: Western Blotting

Background: Tyrosine hydroxylase (TH) catalyzes the rate-limiting step in the synthesis of the neurotransmitter dopamine and other catecholamines. TH functions as a tetramer, with each subunit composed of a regulatory and catalytic domain, and exists in several different isoforms (1,2). This enzyme is required for embryonic development since TH knockout mice die before or at birth (3). Levels of transcription, translation and posttranslational modification regulate TH activity. The amino-terminal regulatory domain contains three serine residues: Ser9, Ser31 and Ser40. Phosphorylation at Ser40 by PKA positively regulates the catalytic activity of TH (4-6). Phosphorylation at Ser31 by CDK5 also increases the catalytic activity of TH through stabilization of TH protein levels (7-9).

$260
100 µl
APPLICATIONS
REACTIVITY
Human, Monkey

Application Methods: Western Blotting

Background: Sphingomyelinases (SMases) catalyze the hydrolysis of sphingomyelin to produce ceramide and phosphocholine (1). Ceramide is an important bioactive lipid triggering signal transduction involved in cell proliferation, apoptosis and differentiation (1,2). A number of SMases have been described and categorized based on their optimum pH activity, cation dependence, tissue distribution, and subcellular localization (1). These include a lysosomal acid SMase, a Zn++-dependent secreted acid SMase, a membrane-bound Mg++-dependent neutral SMase, a Mg++-independent neutral SMase, and an alkaline SMase.

$269
100 µl
APPLICATIONS
REACTIVITY
Human, Mouse, Rat

Application Methods: Immunofluorescence (Frozen), Immunofluorescence (Immunocytochemistry), Immunohistochemistry (Paraffin), Western Blotting

Background: Tyrosine hydroxylase (TH) catalyzes the rate-limiting step in the synthesis of the neurotransmitter dopamine and other catecholamines. TH functions as a tetramer, with each subunit composed of a regulatory and catalytic domain, and exists in several different isoforms (1,2). This enzyme is required for embryonic development since TH knockout mice die before or at birth (3). Levels of transcription, translation and posttranslational modification regulate TH activity. The amino-terminal regulatory domain contains three serine residues: Ser9, Ser31 and Ser40. Phosphorylation at Ser40 by PKA positively regulates the catalytic activity of TH (4-6). Phosphorylation at Ser31 by CDK5 also increases the catalytic activity of TH through stabilization of TH protein levels (7-9).

$260
100 µl
APPLICATIONS
REACTIVITY
Human

Application Methods: Western Blotting

Background: Sphingomyelinases (SMases) catalyze the hydrolysis of sphingomyelin to produce ceramide and phosphocholine (1). Ceramide is an important bioactive lipid triggering signal transduction involved in cell proliferation, apoptosis and differentiation (1,2). A number of SMases have been described and categorized based on their optimum pH activity, cation dependence, tissue distribution, and subcellular localization (1). These include a lysosomal acid SMase, a Zn++-dependent secreted acid SMase, a membrane-bound Mg++-dependent neutral SMase, a Mg++-independent neutral SMase, and an alkaline SMase.

$260
100 µl
APPLICATIONS
REACTIVITY
Human, Mouse, Rat

Application Methods: Immunofluorescence (Frozen), Immunofluorescence (Immunocytochemistry), Western Blotting

Background: Tyrosine hydroxylase (TH) catalyzes the rate-limiting step in the synthesis of the neurotransmitter dopamine and other catecholamines. TH functions as a tetramer, with each subunit composed of a regulatory and catalytic domain, and exists in several different isoforms (1,2). This enzyme is required for embryonic development since TH knockout mice die before or at birth (3). Levels of transcription, translation and posttranslational modification regulate TH activity. The amino-terminal regulatory domain contains three serine residues: Ser9, Ser31 and Ser40. Phosphorylation at Ser40 by PKA positively regulates the catalytic activity of TH (4-6). Phosphorylation at Ser31 by CDK5 also increases the catalytic activity of TH through stabilization of TH protein levels (7-9).

$260
100 µl
APPLICATIONS
REACTIVITY
Hamster, Human, Monkey, Mouse, Rat

Application Methods: Immunofluorescence (Immunocytochemistry), Western Blotting

Background: β-galactosidase (also known as β-gal) is an essential hydrolase enzyme that catalyzes the hydrolysis of galactose-containing carbohydrates into monosaccharides. Substrates of β-galactosides include lactose, various glycoproteins, ganglioside GM1, and lactosylceramides. β-galactosidase is used widely in molecular biology; for example, isolation of recombinant bacteria during molecular cloning utilizes α-complementation of the bacterial β-galactosidase gene (lacZ) in the presence of a β-gal substrate to identify recombinant clones (1). In cell biology, Senescence-Associated beta-galactosidase (SA-β-gal), defined as β-gal activity at pH 6.0, is a widely used marker of replicative senescence. While initially thought to derive from a unique isoform of β-galactosidase expressed specifically in senescent cells (2), SA-β-gal activity was subsequently shown to result from overexpression and accumulation of β-galactosidase in endogenous lysosomes, and is not specifically required for replicative senescence (3).

$108
250 PCR reactions
500 µl
SimpleChIP® Human GLA Promoter Primers contain a mix of forward and reverse PCR primers that are specific to a region of the human alpha-galactosidase A (GLA) promoter. These primers can be used to amplify DNA that has been isolated using chromatin immunoprecipitation (ChIP). Primers have been optimized for use in SYBR® Green quantitative real-time PCR and have been tested in conjunction with SimpleChIP® Enzymatic Chromatin IP Kits #9002 and #9003 and ChIP-validated antibodies from Cell Signaling Technology®.
REACTIVITY
Human

Background: The chromatin immunoprecipitation (ChIP) assay is a powerful and versatile technique used for probing protein-DNA interactions within the natural chromatin context of the cell (1,2). This assay can be used to either identify multiple proteins associated with a specific region of the genome or to identify the many regions of the genome bound by a particular protein (3-6). ChIP can be used to determine the specific order of recruitment of various proteins to a gene promoter or to "measure" the relative amount of a particular histone modification across an entire gene locus (3,4). In addition to histone proteins, the ChIP assay can be used to analyze binding of transcription factors and co-factors, DNA replication factors, and DNA repair proteins. When performing the ChIP assay, cells are first fixed with formaldehyde, a reversible protein-DNA cross-linking agent that "preserves" the protein-DNA interactions occurring in the cell (1,2). Cells are lysed and chromatin is harvested and fragmented using either sonication or enzymatic digestion. Fragmented chromatin is then immunoprecipitated with antibodies specific to a particular protein or histone modification. Any DNA sequences that are associated with the protein or histone modification of interest will co-precipitate as part of the cross-linked chromatin complex and the relative amount of that DNA sequence will be enriched by the immunoselection process. After immunoprecipitation, the protein-DNA cross-links are reversed and the DNA is purified. Standard PCR or quantitative real-time PCR are often used to measure the amount of enrichment of a particular DNA sequence by a protein-specific immunoprecipitation (1,2). Alternatively, the ChIP assay can be combined with genomic tiling micro-array (ChIP on chip) techniques, high throughput sequencing (ChIP-Seq), or cloning strategies, all of which allow for genome-wide analysis of protein-DNA interactions and histone modifications (5-8). SimpleChIP® primers have been optimized for amplification of ChIP-isolated DNA using real-time quantitative PCR and provide important positive and negative controls that can be used to confirm a successful ChIP experiment.