Insulin inactivation

Even though AMP, not cAMP, may be the protein kinase activator, glucagon causes its activation and insulin, inactivation. Details on such hormone effects are lacking. Also recall that malonyl-CoA inhibits palmitoyl-CoA-camitine acyltrans ferase, the rate-controlling enzyme in the /3-oxidation process. Thus, lipid oxidation is inhibited in an environment that favors lipid synthesis, as in the fed state, whereas lipid biosynthesis is inhibited in an environment favoring lipid oxidation, as in fasting. 

Figure 21 20 Insulin inactivates glycogen synthase kinase.

Lipoprotein lipase is activated by high levels of insulin. It acts to extract FAs and glycerol from chylomicrons in the circulation, which are taken up by the adipocytes and re-esterified into TAG to be stored. Insulin inactivates hormone-sensitive lipase to ensure that TAG is not degraded. (Be careful not to get the two enzymes confused.) In the fed state the brain and erythrocytes exclusively use glucose as their fuel supply. Muscle uses glucose as its main fuel, and is also able to metabolize branched amino acids (leucine, isoleucine and valine) as well. 

Increased lipid synthesis/inhibi-tion of lipolysis Activation of lipoprotein lipase (LPL)/induc-tion of fatty acid synthase (FAS)/inactivation of hormone sensitive lipase (HSL) Facilitated uptake of fatty acids by LPL-dependent hydrolysis of triacylglycerol from circulating lipoproteins. Increased lipid synthesis through Akt-mediated FAS-expression. Inhibition of lipolysis by preventing cAMP-dependent activation of HSL (insulin-dependent activation of phosphodiesterases )... 

Also, phosphorylation of Akt results in activation of sterol regulatory-element binding protein 1 (SREBP1), a key transcription factor involved in regulation of lipogenic enzymes. In addition, some of the effects of insulin on cell proliferation and survival may be explained by an Akt-dependent inhibition of apoptosis through phosphorylation and inactivation of proa-poptotic proteins (e.g., BAD, Caspase 9). 

Figure 2. Mechanism of PDH. The three different subunits of the PDH complex in the mitochondrial matrix (E, pyruvate decarboxylase E2, dihydrolipoamide acyltrans-ferase Ej, dihydrolipoamide dehydrogenase) catalyze the oxidative decarboxylation of pyruvate to acetyl-CoA and CO2. E, decarboxylates pyruvate and transfers the acetyl-group to lipoamide. Lipoamide is linked to the group of a lysine residue to E2 to form a flexible chain which rotates between the active sites of E, E2, and E3. E2 then transfers the acetyl-group from lipoamide to CoASH leaving the lipoamide in the reduced form. This in turn is oxidized by E3, which is an NAD-dependent (low potential) flavoprotein, completing the catalytic cycle. PDH activity is controlled in two ways by product inhibition by NADH and acetyl-CoA formed from pyruvate (or by P-oxidation), and by inactivation by phosphorylation of Ej by a specific ATP-de-pendent protein kinase associated with the complex, or activation by dephosphorylation by a specific phosphoprotein phosphatase. The phosphatase is activated by increases in the concentration of Ca in the matrix. The combination of insulin with its cell surface receptor activates PDH by activating the phosphatase by an unknown mechanism.

Both phosphorylase a and phosphorylase kinase a are dephosphorylated and inactivated by protein phos-phatase-1. Protein phosphatase-1 is inhibited by a protein, inhibitor-1, which is active only after it has been phosphorylated by cAMP-dependent protein kinase. Thus, cAMP controls both the activation and inactivation of phosphorylase (Figure 18-6). Insulin reinforces this effect by inhibiting the activation of phosphorylase b. It does this indirectly by increasing uptake of glucose, leading to increased formation of glucose 6-phosphate, which is an inhibitor of phosphorylase kinase. 

Villar-Palasi C On the mechanism of inactivation of muscle glycogen phosphorylase by insulin. Biochim Biophys Acta 1994 1224 384. 

Figure 21-6. Regulation of acetyl-CoA carboxylase by phosphorylation/dephosphorylation.The enzyme is inactivated by phosphorylation by AMP-activated protein kinase (AMPK), which in turn is phosphorylated and activated by AMP-activated protein kinase kinase (AMPKK). Glucagon (and epinephrine), after increasing cAMP, activate this latter enzyme via cAMP-dependent protein kinase. The kinase kinase enzyme is also believed to be activated by acyl-CoA. Insulin activates acetyl-CoA carboxylase, probably through an "activator" protein and an insulin-stimulated protein kinase.

In adipose tissue, the effect of the decrease in insulin and increase in glucagon results in inhibition of lipo-genesis, inactivation of lipoprotein lipase, and activation of hormone-sensitive lipase (Chapter 25). This leads to release of increased amounts of glycerol (a substrate for gluconeogenesis in the liver) and free fatty acids, which are used by skeletal muscle and liver as their preferred metabolic fuels, so sparing glucose. 

DPP-4 is a serine protease that inactivates GLP-1. GLP-1 stimulates insulin secretion and suppresses glucagon release. The inhibition of DPP-4 prolongs the half-life of GLP-1 and brings about beneficial effects on glucose levels and glucose tolerance in type 2 diabetics. Backes et al. [64] report on the parallel optimization of enzyme binding affinity and inhibition, selectivity, ADME properties, and PK (Scheme 19). 

Parenteral administration is not perceived as a problem in the context of drugs which are administered infrequently, or as a once-off dose to a patient. However, in the case of products administered frequently/daily (e.g. insulin to diabetics), non-parenteral delivery routes would be preferred. Such routes would be more convenient, less invasive, less painful and generally would achieve better patient compliance. Alternative potential delivery routes include oral, nasal, transmucosal, transdermal or pulmonary routes. Although such routes have proven possible in the context of many drugs, routine administration of biopharmaceuticals by such means has proven to be technically challenging. Obstacles encountered include their high molecular mass, their susceptibility to enzymatic inactivation and their potential to aggregate. 

Insulin promotes amino acid uptake and protein formation. AKT, noted above, is also implicated in mechanisms which regulate protein synthesis. Acting via GSK-3 again, under basal conditions, GSK-3 phosphorylates a key protein translation regulator (called eIF2B). Thus, if GSK-3 is inactivated, eIF2B is not phosphorylated and mRNA translation is permitted. 

Figure 7.15 Inhibition of acetyl-CoA carboxylase by cyclic AMP dependent protein kinase and AMP dependent protein kinase the dual effect of glucagon. Phosphorylation of acetyl-CoA carboxylase by either or both enzymes inactivates the enzyme which leads to a decrease in concentration of malonyl-CoA, and hence an increase in activity of carnitine palmitoyltransferase-I and hence an increase in fatty acid oxidation. Insulin decreases the cyclic AMP concentration maintaining an active carboxylase and a high level of malonyl-CoA to inhibit fatty acid oxidation.

Unwanted effects. Hypoglycemia results from absolute or relative overdosage (see p. 260). Allergic reactions are rare—locally redness at injection site, atrophy of adipose tissue (lipodystrophy) systemically urticaria, skin rash, anaphylaxis. Insulin resistance can result from binding to inactivating antibodies. A possible local lipohypertrophy can be avoided by alternating injection sites. 

In contrast to glucagon, the peptide hormone insulin (see p. 76) increases glycogen synthesis and inhibits glycogen breakdown. Via several intermediates, it inhibits protein kinase GSK-3 (bottom right for details, see p. 388) and thereby prevents inactivation of glycogen synthase. In addition, insulin reduces the cAMP level by activating cAMP phosphodiesterase (PDE). 

Interconversion processes (see p. 120) also play an important role. They are shown here in detail using the example of the PDH complex (see p. 134). The inactivating protein kinase [la] is inhibited by the substrate pyruvate and is activated by the products acetyl-CoA and NADH+H. The protein phosphatase [Ibj—like isodtrate dehydrogenase [3] and the ODH complex [4j-is activated by Ca. This is particularly important during muscle contraction, when large amounts of ATP are needed. Insulin also activates the PDH complex (through inhibition of phosphorylation) and thereby promotes the breakdown of glucose and its conversion into fatty acids. 

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