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Mitochondrial Dysfunction and Metabolic Defects

BioFiles 2011, 6.4, 14.

Defects in mitochondrial function are associated with insulin resistance in both human and animal studies. Insulin resistance is associated with a decrease in the number, oxidative capacity, size, and density of mitochondria. Understanding the mechanisms that lead to decreased activity and/or biogenesis of mitochondria in insulin-sensitive tissues is therefore of vital importance to developing therapies to reverse insulin resistance.

Exposure to high levels of lipids is associated with an increased accumulation of intracellular fatty acids and decreases in mitochondrial function in liver and skeletal muscle tissue. Increased exposure of cells to fatty acids may lead to the accumulation of toxic proinflammatory lipid components that activate stress kinase signaling pathways resulting in the antagonism of insulin receptor signaling. Alternatively, increased rates of fatty acid metabolism may exceed the capacity of the tricarboxylic acid (TCA) cycle and electron transport chain resuling in incomplete fatty acid oxidation and the production of toxic metabolites.

Mitochondrial Membrane

Once transported across the mitochondrial membrane, fatty acids are metabolized through beta oxidation releasing acetyl-CoA molecules. Acetyl CoA enters the TCA cycle resulting in the production of NADH and FADH2. NADH and FADH2 are used by the electron transport chain to produce ATP.


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Key Metabolites Involved in Fatty Acid Metabolism


Name Description Catalog No.
Acetyl coenzyme A sodium salt An essential cofactor in enzymatic acetyl transfer reactions. Acetyl-CoA is an essential cofactor and carrier of acyl groups in enzymatic acetyl transfer reactions. It is formed either by the oxidative decarboxylation of pyruvate in mitochondria, by the oxidation of long-chain fatty acids, or by the oxidative degradation of certain amino acids. Acetyl-CoA is the starting compound for the citric acid cycle (Kreb's cycle). It is also a key precursor in lipid biosynthesis, and the source of all fatty acid carbons. Acetyl-CoA positively regulates the activity pyruvate carboxylase. It is a precursor of the neurotransmitter acetylcholine. Histone acetylases (HAT) use Acetyl-CoA as the donor for the acetyl group use in the post-translational acetylation reactions of histone and non-histone proteins. A2056-1MG
A2056-5MG
A2056-10MG
A2056-25MG
A2056-100MG
Acetyl coenzyme A trilithium salt An essential cofactor in enzymatic acetyl transfer reactions. Acetyl-CoA is an essential cofactor and carrier of acyl groups in enzymatic acetyl transfer reactions. It is formed either by the oxidative decarboxylation of pyruvate in mitochondria, by the oxidation of long-chain fatty acids, or by the oxidative degradation of certain amino acids. Acetyl-CoA is the starting compound for the citric acid cycle (Kreb's cycle). It is also a key precursor in lipid biosynthesis, and the source of all fatty acid carbons. Acetyl-CoA positively regulates the activity pyruvate carboxylase. It is a precursor of the neurotransmitter acetylcholine. Histone acetylases (HAT) use Acetyl-CoA as the donor for the acetyl group use in the post-translational acetylation reactions of histone and non-histone proteins. A2181-1MG
A2181-5MG
A2181-10MG
A2181-25MG
A2181-100MG
Butyryl coenzyme A lithium salt hydrate Butyryl CoA is a substrate for Butyryl CoA Dehydrogenase. Butyryl CoA is involved in both lipid and butanoate metabolism. B1508-5MG
B1508-10MG
B1508-25MG
L-Carnitine hydrochloride Carnitine is a quaternary amine that occurs naturally in most mammalian tissue. It is present in relatively high concentrations in skeletal muscle and heart where it is involved in regulating energy metabolism. It shifts glucose metabolism from glycolysis to glycogen storage and enhances the transport of long chain fatty acids into the mitochondria where they are oxidized for energy production. C0283-10MG
C0283-1G
C0283-5G
C0283-25G
C0283-100G
Decanoyl coenzyme A monohydrate D5269-5MG
D5269-25MG
n-Heptadecanoyl coenzyme A lithium salt Heptadecanoyl CoA is a lipid metabolism intermediate formed by the covalent attachment of Heptadecanoic acid to Coenzyme A. H1385-5MG
DL-Hexanoylcarnitine chloride H2132-25MG
Hexanoyl coenzyme A trilithium salt hydrate Hexanoyl CoA is involved in fatty acid oxidation, lipid biosynthesis, and ceramide formation. H2012-5MG
H2012-10MG
Lauroyl coenzyme A lithium salt Lauroyl CoA is a substrate for Protein FAM 34A. Lauroyl CoA is involved in lipid biosynthesis and fatty acid transport. L2659-5MG
L2659-25MG
Malonyl coenzyme A lithium salt Malonyl Coenzyme A is a coenzyme A derivative that is utilized in fatty acid and polyketide synthesis and in the transport of a-ketoglutarate across the mitochondrial membrane. Malonyl CoA is formed by the Acetyl CoA Carboxylase-mediated carboxylation of acetyl CoA. M4263-5MG
M4263-10MG
M4263-25MG
M4263-100MG
Malonyl coenzyme A tetralithium salt Malonyl Coenzyme A is a Coenzyme A derivative that is utilized in fatty acid and polyketide synthesis and in the transport of a-ketoglutarate across the mitochondrial membrane. Malonyl CoA is formed by the Acetyl CoA Carboxylase-mediated carboxylation of acetyl CoA. 63410-10MG-F
63410-50MG-F
Myristoyl-dl-carnitine chloride M0135-100MG
Myristoyl coenzyme A lithium salt M4414-5MG
Octanoyl coenzyme A lithium salt hydrate Medium-chain fatty acid covalently linked to coenzyme A. O6877-5MG
O6877-10MG
O6877-25MG
Palmitoyl-L-carnitine chloride Long-chain acylcarnitine and well-known intermediate in mitochondrial fatty acid oxidation. Modifies myocardial levels of high-energy phosphates and free fatty acids in the heart. Increases erythroid colony formation in culture. Reduces surface negative charge of erythrocytes and myocytes. Reported to affect currents and inhibit endothelium-dependent relaxation induced by acetylcholine and substance P in a dose-dependent manner by suppressing the intracellular calcium signal transduction in endothelial cells. Inhibits the Na/K pump current but has no effect on the intracellular calcium current in guinea pig ventricular cells. However, like oubain, it reversibly depolarizes the resting membrane, decreases action potential duration, and increases the amplitude of myocyte contractions. P1645-5MG
P1645-10MG
P1645-25MG
Palmitoyl coenzyme A lithium salt Long-chain fatty acid (C16) covalently linked to coenzyme A. P9716-5MG
P9716-10MG
P9716-25MG
P9716-100MG
n-Propionyl coenzyme A lithium salt Propionyl coenzyme A (CoA) is the coenzyme A derivative of propionic acid. Propionyl CoA is formed during the ß-oxidation of odd-chain fatty acids. Propionyl CoA is also formed during the metabolism of isoleucine and valine. P5397-5MG
P5397-10MG
P5397-25MG
Sodium citrate tribasic dihydrate C7254-1KG
C7254-5KG
C7254-10KG
Stearoyl coenzyme A lithium salt Stearoyl CoA is a saturated fatty acid metabolite involved in polyunsaturated fatty acid synthesis and the PPAR signaling pathway. S0802-5MG
S0802-10MG
S0802-25MG
Succinyl coenzyme A sodium salt Succinyl CoA is an intermediate in the citric acid cycle. It is formed by α-ketoglutarate dehydrogenase by the decarboxylation of a-ketoglutarate. Succinyl CoA is also formed from propionyl CoA during the ß-oxidation of odd-chain fatty acids. Succinyl CoA serves as a precursor in heme synthesis. It is also required for the oxidation of ketone bodies. S1129-5MG
S1129-25MG

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Enzymes that Regulate Fatty Acid Metabolism


Name Description Catalog No.
Acetyl-CoA Carboxylase 1 human Acetyl-CoA Carboxylase (ACC) regulates the metabolism of fatty acids. This enzyme catalzes the formation of Malonyl CoA through the irreversible carboxylation of acetyl CoA. There are two main isoforms of Acetyl-CoA carboxylase expressed in mammals, Acetyl-CoA carboxylase 1 (ACACA) and Acetyl-CoA carboxylase 2 (ACACB). ACACA has broad tissue distribution but is enriched in tissues critical for fatty acid sythesis such as adipose tissue. ACACB is enriched in tissues such as skeletal muscle and heart that are critical for fatty acid oxidation.

The Acetyl-CoA Carboxylase enzymes are activated by citrate, glutamate, and dicarboxylic acids and negatively regulated by long and short chain fatty acyl CoAs. Acetyl-CoA Carboxylase 1 is essential for breast cancer and prostrate cancer cell survival. Because of thier roles in fatty acid metabolism and oxidation, ACACA and ACACB are therapeutic targets for treating obesity and metabolic syndrome disorders.
A6986-10UG
Acetyl-CoA carboxylase 2 human Acetyl-CoA Carboxylase (ACC) regulates the metabolism of fatty acids. This enzyme catalzes the formation of Malonyl CoA through the irreversible carboxylation of acetyl CoA. There are two main isoforms of Acetyl-CoA carboxylase expressed in mammals, Acetyl-CoA carboxylase 1 (ACACA) and Acetyl-CoA carboxylase 2 (ACACB). ACACA has broad tissue distribution but is enriched in tissues critical for fatty acid sythesis such as adipose tissue. ACACB is enriched in tissues such as skeletal muscle and heart that are critical for fatty acid oxidation.

The Acetyl-CoA Carboxylase enzymes are activated by citrate, glutamate, and dicarboxylic acids and negatively regulated by long and short chain fatty acyl CoAs. Because of thier roles in fatty acid metabolism and oxidation, ACACA and ACACB are therapeutic targets for treating obesity and metabolic syndrome disorders.
A6861-10UG
SIRT1 human SIRT1 plays a pivotal role in the regulation of cellular differentiation, metabolism, cell cycle, apoptosis and regulation of p53. S8446-150UG
AMPK (A1/B1/G1), active, His tagged human AMPK is a heterotrimer protein kinase consisting of a a catalytic subunit, and non-catalytic ß and γ subunits. AMPK is an important energy-sensing enzyme that monitors cellular energy status. In response to cellular metabolic stresses, AMPK is activated and phosphorylates and inactivates acetyl-CoA carboxylase (ACC) and 3-hydroxy-3-methylglutaryl-CoA reductase (HMGCR), key enzymes involved in regulating biosynthesis of fatty acid and cholesterol. A1233-10UG
AMPK (A1/B1/G2), active, His tagged human AMP-activated protein kinase (AMPK) exhibits a key role as a master regulator of cellular energy homeostasis. AMPK exists as a heterotrimeric complex composed of a catalytic a subunit and regulatory ß and γ subunits. Binding of AMP to the γ subunit allosterically activates the complex. AMPK is activated in response to stresses that deplete cellular ATP (low glucose, hypoxia and ischemia) and via signaling pathways in response to adiponectin, leptin and CAMKKb. A1358-10UG
AMPK (A2/B1/G1), active, His tagged human AMPK (A2/B1/G1) plays a key role in insulin signaling pathway and is a major therapeutic target for the treatment of diabetes. AMPK is viewed as a fuel sensor for glucose and lipid metabolism by modulating the activity of the autonomous nervous system in vivo. Short-term overexpression of a constitutively active form of AMPK in the liver leads to mild hypoglycemia and fatty liver due to increased fatty acid utilization. A1733-10UG
AMPK (A1/B1/G3), active, His tagged human AMPK (A1/B1/G3) is a member of the AMPK family which are heterotrimeric proteins consisting of an a catalytic subunit, and non-catalytic ß and γ subunits. AMPKs are an important energy-sensing enzyme group in the cells that monitor energy status particularly in response to stress. AMPKs regulate fatty acid and cholesterol synthesis by regulating the key rate-limiting enzymes acetyl-CoA carboxylase and 3-hydroxy-3-methylglutaryl- CoA reductase. The γ subunit is dominantly expressed in skeletal muscle where it may play a key role in the regulation of energy metabolism. A4486-10UG
AMPK (A1/B2/G1), active, His tagged human AMPK (A1/B2/G1) is a member of the AMPK family which are heterotrimeric proteins consisting of an a catalytic subunit, and non-catalytic ß and γ subunits. AMPKs are an important energy-sensing enzyme group in the cells that monitor energy status particularly in response to stress. AMPKs regulate fatty acid and cholesterol synthesis by regulating the key rate-limiting enzymes acetyl-CoA carboxylase and 3-hydroxy-3-methylglutaryl- CoA reductase. The ß subunit may be a positive regulator of AMPK activity and is highly expressed in skeletal muscle. A4611-10UG
GSK3ß, active, His tagged human GSK3ß is a serine threonine protein kinase that was originally identified as the kinase that phosphorylates and inhibits glycogen synthase.GSK3ß is ubiquitously present in human tissues and implicated in the regulation of several physiological processes, including the control of glycogen and protein synthesis by insulin and modulation of the transcription factors AP-1 and CREB. Transient transfection of human GSK3ß into Chinese hamster ovary cells stably transfected with individual human tau isoforms leads to hyperphosphorylation of tau at all the sites investigated with phosphorylation-dependent anti-tau antibodies. G4296-10UG

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Antibodies Against Key Fatty Acid Metabolism Proteins


Name Host Reacts With Application Prestige
Antibody
Catalog No.
Anti-ABAD/HADH2 goat Xenopus
canine
chimpanzee
human
mouse
rat
zebrafish
ELISA (i)
WB
- SAB2500007-100UG
Anti-ACAA1 rabbit human WB - SAB2100018-50UG
Anti-ACAA1 rabbit human IF (i)
IHC (p)
PA
WB
Yes HPA007244-100UL
Anti-ACAA1 rabbit human IHC (p)
PA
WB
Yes HPA006764-100UL
Monoclonal Anti-ACAA1 mouse human ELISA (i)
IHC (p)
WB
- SAB1402104-100UG
Anti-ACAA2 rabbit human WB - SAB2100019-50UG
Monoclonal Anti-ACAA2 mouse human
mouse
rat
ELISA (i)
IF (i)
IHC (p)
WB
- WH0010449M1-100UG
Anti-ACAD8 (ab1) chicken human WB - GW21999A-50UG
Anti-ACAD9 mouse human ELISA (c)
WB
- SAB1400510-50UG
Anti-ACAD9 rabbit human IHC (p)
PA
Yes HPA037716-100UL
Anti-ACAD9 (76–89) rabbit human IF (i)
WB
- SAB1101278-200UL
Anti-ACAD9 (406–420) rabbit human IF (i)
WB
- SAB1101279-200UL
Anti-ACAD-8 (ab2) chicken human WB - GW21999B-50UG
Anti-ACADL rabbit human WB - AV33855-50UG
Anti-ACADL rabbit human WB - SAB2100020-50UG
Anti-ACADL rabbit human IHC (p)
PA
WB
Yes HPA011990-100UL
Anti-ACADL rabbit human IF (i)
IHC (p)
PA
Yes HPA010611-100UL
Anti-ACADM goat human ELISA (i)
WB
- SAB2500017-100UG
Anti-ACADM rabbit human IF (i)
IHC (p)
PA
WB
Yes HPA006198-100UL
Anti-ACADM rabbit human WB - AV32789-100UG
Anti-ACADM rabbit human IHC (p)
PA
WB
Yes HPA026542-100UL
Anti-ACADM rabbit human WB - AV32788-50UG
Monoclonal Anti-ACADM, (N-terminal) mouse human ELISA (i) - SAB1402105-100UG
Anti-ACADS mouse human ELISA (c)
WB
- SAB1400001-50UG
Anti-ACADS rabbit human WB - SAB2100021-50UG
Anti-ACADS rabbit human IHC (p)
PA
Yes HPA004799-100UL
Anti-ACADS rabbit human IHC (p)
PA
WB
Yes HPA022271-100UL
Anti-ACADS (376–390) rabbit human WB - A4984-200UL
Anti-ACADSB (316–330) rabbit human WB - A5109-200UL
Anti-ACADSB (396–410) rabbit human WB - A5234-200UL
Anti-ACADVL rabbit human IHC (p)
PA
WB
Yes HPA020595-100UL
Anti-ACADVL rabbit human WB - AV54486-50UG
Anti-ACADVL rabbit human IHC (p)
PA
WB
Yes HPA019006-100UL
Monoclonal Anti-ACADVL mouse human ELISA (i)
IHC (p)
WB
- WH0000037M1-100UG
Anti-ACADVL (386–400) rabbit human WB - A5484-200UL
Anti-ACAT1 rabbit human
mouse
rat
ELISA (i)
WB
- SAB4501646-100UG
Anti-ACAT1 rabbit human WB - AV54278-50UG
Anti-ACAT1 rabbit human IF (i)
IHC (p)
PA
WB
Yes HPA004428-100UL
Anti-ACAT1 rabbit human IF (i)
IHC (p)
PA
WB
Yes HPA007569-100UL
Anti-ACAT2 rabbit human IF (i)
IHC (p)
PA
WB
Yes HPA025765-100UL
Anti-ACAT2 rabbit human IF (i)
IHC (p)
PA
WB
Yes HPA025736-100UL
Anti-ACAT2 rabbit human IHC (p)
PA
WB
Yes HPA025811-100UL
Monoclonal Anti-ACAT2 mouse human ELISA (c)
ELISA (i)
WB
- SAB1402106-100UG
Anti-ACAT2 (216-230) rabbit human WB - A5609-200UL
Anti-ACAT2 (AB1) rabbit human IHC
WB
- AV32790-100UG
Anti-ACAT2 (AB2) rabbit human IHC
WB
- AV32791-100UG
Anti-Acetyl-CoA Acetyltransferase 2 chicken human WB - GW22016-50UG
Anti-ACADS rabbit human
mouse
rat
WB - SAB2103528-50UG
Anti-EHHADH mouse human ELISA (c)
IF (i)
WB
- SAB1400088-50UG
Anti-EHHADH rabbit human WB - SAB1401124-100UG
Anti-EHHADH rabbit human
rat
IHC
WB
- SAB4500727-100UG
Anti-EHHADH rabbit human IHC (p)
PA
WB
Yes HPA036401-100UL
Anti-EHHADH (346–360) rabbit human IF (i)
WB
- E4534-200UL
Anti-ERAB rabbit human
mouse
rat
IHC
WB
- SAB4501374-100UG
Anti-ERAB rabbit human IHC (p)
WB
- E1151-25UG
Anti-HADH rabbit human WB - SAB2101011-50UG
Anti-HADH rabbit human IHC (p)
PA
WB
Yes HPA039588-100UL
Anti-HADH (81-95) rabbit human WB - SAB1100024-200UL
Anti-HADH (300-314) rabbit human WB - SAB1100025-200UL
Anti-HADHA rabbit human IF (i)
IHC (p)
PA
WB
Yes HPA015536-100UL
Anti-HADHB rabbit human IHC (p)
PA
WB
Yes HPA037539-100UL
Anti-HADHB rabbit human IHC
WB
- AV48133-50UG
Anti-HADH/HADHSC goat canine
human
mouse
rat
ELISA (i)
WB
- SAB2500501-100UG
Anti-HADHSC chicken human
mouse
rat
WB - GW22261-50UG
Monoclonal Anti-HADHSC mouse human ELISA (i)
IF (i)
IHC (p)
WB
- WH0003033M1-100UG
Anti-HSD17B10 rabbit human IHC (p)
PA
WB
Yes HPA001432-100UL
Anti-HSD17B4 rabbit human IHC (p)
PA
WB
Yes HPA021311-100UL
Anti-HSD17B4 rabbit human IHC (p)
PA
WB
Yes HPA021302-100UL
Anti-HSD17B4 rabbit human IHC (p)
PA
WB
Yes HPA021479-100UL
Anti-HSD17B4 (131-145) rabbit human WB - SAB1100185-200UL
Anti-HSD17B4 (572-585) rabbit human WB - SAB1100186-200UL

Immunofluorescence
Anti-ACAA1: Cat. No. HPA007244: Immunofluorescent staining of human cell line U-251MG shows positivity in vesicles.


Immunofluorescence
Monoclonal Anti-ACAA2, Cat. No. WH0010449M1; antibody concentration: 40 µg/mL using HeLa cell.


Immunofluorescence
Anti-ACADL: Cat. No. HPA010611: Immunofluorescent staining of human cell line U-2 OS shows positivity in cytoplasm and mitochondria.


Immunofluorescence
Anti-ACAT1: Cat. No. HPA004428: Immunofluorescent staining of human cell line A-431 shows positivity in mitochondria.


Immunofluorescence
Anti-EHHADH (346-360) : Cat. No. E4534: Immunofluorescence of HUVEC cells using EHHADH (346-360) , Cat. No. E4534 (red) at a 1:50 dilution, taken at 40x magnification and nuclear staining with Hoescht 33342 (blue).Yale HTCB IF procedure used. Images have been adjusted to improve viewing quality. If you would like the original image, please send a request to protocols@sial.com.


Immunohistochemistry
Anti-HADH: Cat. No. HPA039588: Immunohistochemical staining of human liver shows strong cytoplasmic positivity in hepatocytes.


Immunohistochemistry
Anti-HSD17B4: Cat. No. HPA021302: Immunohistochemical staining of human liver shows strong cytoplasmic positivity in hepatocytes.

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Materials

     
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