Carbohydrate metabolism

BASIC SCIENCE

Carbohydrate metabolism

carbohydrate per day. About 80e100 g is stored in the liver, from where it can be released into the blood for transport to other tissues. The largest carbohydrate store is in skeletal muscle, but the muscle glycogen reserve is not readily available to other tissues. The glucose content of extracellular fluid amounts to only about 15 g, and should not be regarded as an energy store, as even small decreases in blood glucose concentration can impair muscle and nerve cell function. The important dietary carbohydrates consist of monosaccharides (such as glucose, fructose and galactose), disaccharides (pairs of sugar molecules linked together, such as maltose, sucrose and lactose) and polysaccharides (such as starch, which consists of long chains of glucose molecules) (Figure 1). Sucrose and starch are normally the main carbohydrates in the diet, and varying the proportions of these has important nutritional and health implications. Dietary carbohydrates are digested to their component monosaccharides by hydrolysis of the bonds linking the monosaccharides before absorption occurs. Some hydrolysis occurs in the mouth and stomach, but most occurs in the upper part of the small intestine, where the pH allows high activity of the specific enzymes secreted into the intestinal lumen. Some polysaccharides, such as cellulose, are resistant to hydrolysis in the human intestine, and pass through the gut largely undigested. If dietary carbohydrate intake exceeds storage capacity, most of the excess is oxidized to provide energy. If, however, the intake of carbohydrate is very high e close to or above total energy demand e some of the excess is converted to fatty acids for storage in the adipose cells.

Ron Maughan

Abstract Carbohydrate normally accounts for about 50% of total dietary energy intake, but the general recommendation is for an increased consumption of complex carbohydrates. After digestion and absorption, carbohydrate is metabolized to provide energy (4 kcal/g) or is stored in muscle and liver as glycogen. The body’s carbohydrate stores are normally about 400e500 g in the fed state. Six-carbon glucose molecules are degraded by a series of chemical reactions to three-carbon pyruvate by the reactions of glycolysis; pyruvate can be further metabolized to lactate. These reactions occur in the cell cytoplasm without the involvement of molecular oxygen, so are described as anaerobic. Pyruvate (and lactate) can be further oxidized to CO2 and water by the reactions of the Krebs’ (tricarboxylic acid) cycle that occur in the mitochondria. Glucose is an essential fuel for the brain and for some other cells, notably red blood cells. Because body carbohydrate reserves are limited, and also because stored fatty acids cannot be converted to carbohydrate, the metabolism of carbohydrates in different tissues is tightly regulated. Some de novo synthesis of glucose is possible from non-carbohydrate sources, including glycerol and the carbon skeletons of some amino acids. Excess dietary carbohydrate is generally oxidized rather than stored.

Keywords Carbohydrate; energy; gluconeogenesis; glycogen; glycolysis; metabolism; sugar

The reactions of anaerobic glycolysis and glycogenolysis Carbohydrate is an important component of the diet, providing energy but also contributing to the taste and texture of foods. Many high-carbohydrate foods are also important sources of vitamins, minerals and other micronutrients. It is usually recommended that carbohydrate should contribute about 50e60% of the daily energy intake, but data from the UK Household Survey suggest that total carbohydrate provides on average about 48% of food energy intake for men and for women. There are no significant age differences for men or women in the proportion of food energy derived from total carbohydrate, but there are very large variations between individuals. At the lower 2.5th percentile, just over one-third of food energy comes from carbohydrate, but at the upper 2.5th percentile carbohydrate contributes close to 60%. For the average Western diet with slightly less than 50% of total energy in the form of carbohydrate, the total daily carbohydrate intake is normally about 200e300 g (800e1200 kcal or 3200e4800 kJ). Carbohydrate is also available to the body from endogenous reserves in the form of muscle and liver glycogen, but these stores are small and may be less than the daily turnover. In a typical 70-kg man, the total body carbohydrate store is about 300e 500 g, but some endurance athletes consume more than 1000 g

The initial steps in the degradation of carbohydrate occur without the involvement of oxygen, and are therefore termed anaerobic processes. Degradation of glucose is referred to as glycolysis, whereas glycogenolysis begins with glycogen. Glycolysis effectively converts a six-carbon glucose molecule to two three-carbon pyruvate molecules (Figure 2). Glycolysis allows some of the chemical energy liberated by the breaking of chemical bonds in the glucose molecule to be conserved in the form of adenosine triphosphate (ATP), which can be used by the cell to power other metabolic processes. A specific transporter protein (GLUT 4) carries glucose across the cell membrane, where a phosphate group is transferred from ATP to form glucose-6-phosphate. This irreversible reaction, catalysed by hexokinase, prevents loss of this valuable nutrient from the cell because phosphorylated sugars cannot cross cell membranes. This also ensures a concentration gradient for glucose transport across the cell membrane. In liver, a phosphatase enzyme catalyses the reverse reaction, allowing free glucose to leave the cell; however, this enzyme is absent from muscle. If glycogen, rather than blood glucose, is the starting point, the first step is to split off a single glucose molecule in a reaction catalysed by glycogen phosphorylase to form glucose-1phosphate and a glycogen molecule that is one glucose residue shorter than the original. The substrates are glycogen and inorganic phosphate; unlike the hexokinase reaction, there is no breakdown of ATP in this reaction. Glucose-1-phosphate is

Ron Maughan BSc (Physiology) PhD is Professor of Sport and Exercise Nutrition at Loughborough University, Loughborough, UK. Conflicts of interest: none declared.

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Structure and classification of important dietary carbohydrates and glycogen

6 CH2OH O

O

H

5 H

O

OH 3

H

6 HOCH2 5

1 2

H

OH

OH

2 H

H 4

HO 3

α-1,4 glycosidic bonds

CH2OH 1

OH

H

OH H Fructose C6H12O6

Glucose C6H12O6 Monosaccharides

6 CH2OH H

H

4 HO

1

O

5 H

3 H

H

H

2

1 (α) OH

O

HOCH2 O

2 OH

α-1,6 glycosidic bonds

5 H

HO

3 OH

4

CH2OH 6

H

Sucrose C12H22O11 Polysaccharide (Glycogen)

Disaccharide

Figure 1

rapidly converted to glucose-6-phosphate, which then proceeds down the glycolytic pathway. Glucose-6-phosphate is converted to fructose-6-phosphate and a second phosphorylation step follows, converting fructose-6phosphate to fructose-1,6-diphosphate. This reaction also requires a phosphate group from ATP, and is catalysed by phosphofructokinase (PFK). The activity of this complex enzyme is affected by many intracellular factors, and it plays an important role in controlling flux through the pathway. The PFK reaction is the first opportunity for regulation at a point that affects the metabolism of both glucose and glycogen. In any sequence of metabolic reactions, there can only be one reaction that determines the overall rate at which the sequence can proceed; it can go no faster than the slowest, or rate-limiting, reaction, and PFK is normally the point in glycolysis that determines the rate of the overall reaction. So far, glycolysis, which is intended to make energy available to the cell, has required the investment of two ATP molecules if glucose was the starting point, or one ATP if glycogen was the starting point, with no immediate return. Fructose-1,6-diphosphate is split into two three-carbon molecules: glyceraldehyde-3-phosphate and dihydroxyacetone phosphate. These two molecules are interconvertible, and further metabolism occurs only through glyceraldehyde-3-phosphate. Each of the succeeding steps in glycolysis thus occurs in duplicate. Glyceraldehyde-3-phosphate is converted to 1,3-diphosphoglyceric acid in a reaction catalysed by glyceraldehyde-3-phosphate

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dehydrogenase and involving the simultaneous conversion of the oxidized form of nicotinamide adenine dinucleotide (NADþ) to its reduced form, NADH. At physiological pH, 1,3-diphosphoglyceric acid exists as the ionized form, 1,3-diphosphoglycerate (1,3DPG), so a hydrogen ion is released. As well as accepting a hydrogen ion, NADþ also accepts two electrons. The additional phosphate group incorporated into 1,3-DPG is derived from inorganic phosphate, so there is no further input of ATP. In the next step, a phosphate group is transferred from 1,3DPG to adenosine diphosphate (ADP), with ATP being formed and the 1,3-DPG converted to 3-phosphoglycerate. Internal reorganization of the 3-phosphoglycerate molecule shifts the phosphate group to the 2 carbon position, to form 2-phosphoglycerate. A dehydration reaction then occurs, catalysed by enolase, which results in the formation of phosphoenolpyruvate. The last step in glycolysis results in the transfer of the phosphate group from phosphoenolpyruvate to ADP, with the formation of pyruvate and another ATP. The net effect of glycolysis is thus conversion of one molecule of glucose to two molecules of pyruvate, with the formation of two molecules of ATP and the conversion of two molecules of NADþ to NADH. If glycogen rather than glucose is the starting point, three molecules of ATP are produced, as there is no initial investment of ATP in the first phosphorylation step. Although this net energy yield appears to be small, the relatively large carbohydrate store available and the rapid rate at which glycolysis can proceed mean that the energy that can be supplied in this

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way is crucial, especially in situations of suddenly increased energy demand such as occurs at the onset of exercise. These reactions can also occur in the absence of oxygen. The reactions of glycolysis occur in the cytoplasm, and the pyruvate formed is not phosphorylated and so can leave the cell. Some pyruvate escapes from tissues such as muscle when the rate of glycolysis is high, but most is further metabolized. The fate of the pyruvate produced by glycolysis depends on a number of factors, including the metabolic capacity of the tissue.

Glycolysis Glycogen

Glucose

ATP

Pi glycogen phosphorylase

hexokinase*

ADP phosphoglucomutase Glucose-6-phosphate Glucose-1-phosphate

glucosephosphate isomerase

Regeneration of NADD When glycolysis proceeds rapidly, the problem for the cell is that the availability of NADþ, which is necessary as a co-factor in the glyceraldehyde-3-phosphate dehydrogenase reaction, becomes limiting. The amount of NADþ in the cell is very small e only about 0.8 mmol per g of muscle e relative to the rate at which glycolysis can proceed. In brief bursts of activity induced by electrical stimulation, muscle ATP turnover can reach 150 mmol/ g/minute. If the NADH formed by glycolysis is not re-oxidized to NADþ at an equal rate, glycolysis will be unable to proceed and to contribute to energy supply. Two processes are available to most (but not all) cells by which oxidation of NADH and regeneration of NADþ can occur. Reduction of pyruvate to lactate (Figure 3) achieves this, and this reaction can proceed in the absence of oxygen. The normal pH of the muscle cell at rest is about 7.1, but this can fall to 6.5 or less in high-intensity exercise or when the oxygen supply is interrupted. The negative effects of the acidosis resulting from lactate accumulation are often stressed, but the energy made available by anaerobic glycolysis allows the generation of ATP that would not otherwise be possible. Alternatively, pyruvate may undergo oxidative metabolism to CO2 and water within the mitochondrion. The first step is the conversion, by oxidative decarboxylation, of the three-carbon pyruvate to a two-carbon acetate group that is linked to coenzyme A to form acetyl-CoA. This reaction, in which NADþ is converted to NADH, is catalysed by the pyruvate dehydrogenase enzyme complex. Acetyl-CoA is oxidized to CO2 and water in the tricarboxylic acid (TCA) cycle; this series of reactions is also

Fructose-6-phosphate

ATP

6-phosphofructokinase*

ADP Fructose-1,6-disphosphate

aldolase

Dihydroxyacetone Glyceraldehyde-3-phosphate phosphate triosephosphate isomerase P i, NAD +

glyceraldehyde-3-phosphate dehydrogenase

NADH

1,3-Diphosphoglycerate

ADP phosphoglycerate kinase ATP 3-Phosphoglycerate

phosphoglyceromutase 2-Phosphoglycerate

enolase

Lactate dehydrogenase reaction Phosphoenolpyruvate

CH3CO COO– +NADH +H+

ADP pyruvate kinase*

ATP

Pyruvate

lactate dehydrogenase Lactate

low pH low NAD+/high NADH

NADH

NAD+

NAD+

NADH

CoA pyruvate dehydrogenase*

LDH

High pH High NAD+/low NADH

CH3CH(OH) COO– + NAD+

Lactate

Acetyl-CoA + CO2 The lactate dehydrogenase reaction catalyses the interconversion of lactate and pyruvate. It can proceed in either direction, depending on substrate concentrations and the ionic environment. LDH, lactate dehydrogenase; NAD, nicotinamide adenine dinucleotide.

Asterisks refer to possible control points where flux through the pathway can be regulated. ADP, adenosine diphosphate; ATP, adenosine triphosphate; CoA, coenzyme A; NAD, nicotinamide adenine dinucleotide; Pi, inorganic phosphate.

Figure 3

Figure 2

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Pyruvate

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BASIC SCIENCE

known as the Krebs’ cycle, after Hans Krebs, or the citric acid cycle, as citrate is one of the key reaction intermediates. The reactions involve combination of acetyl-CoA with oxaloacetate to form citrate, a six-carbon TCA. A series of reactions leads to the sequential loss of hydrogen ions and CO2, resulting in the regeneration of oxaloacetate. Acetyl-CoA is also an intermediate of fatty acid oxidation, and the final steps of oxidative degradation are therefore common to both fat and carbohydrate. The overall reaction starting with glucose as the fuel can be summarized as follows:

metabolism. A high rate of fatty acid oxidation results in citrate accumulation within the mitochondrion; some of this is transported into the cytoplasm, resulting in a reduction in the rate of glycolysis. Inhibition of PFK also causes accumulation of glucose-6-phosphate, which inhibits the activity of hexokinase and so reduces the entry into the cell of glucose that is not needed. Pyruvate kinase activity is regulated by some of the same factors that affect PFK activity, including activation by high ADP concentrations and inhibition by ATP. Pyruvate dehydrogenase is not a single enzyme, but is a complex of three enzymes that can exist in an active (a) dephosphorylated form and an inactive (b) phosphorylated form. Control of the activity of this enzyme complex is central to the integration of fat and carbohydrate metabolism, but the control mechanisms are not well understood.

C6 H12 O6 þ 6O2 þ 38ADP þ 38Pi /6CO2 þ 6H2 O þ 38ATP where C6H12O6 is glucose and Pi is inorganic phosphate. The reactions of oxidative phosphorylation occur within the mitochondria, whereas glycolysis is a cytosolic process, and the inner mitochondrial membrane is impermeable to NADH and to NADþ. A number of substrate shuttles act to transfer reducing equivalents into the mitochondrion. Some of the pyruvate formed may be converted to the amino acid alanine, which can be released from the muscle and transport amino groups to other tissues such as liver and kidney. This is a way of shuttling both three-carbon units and amino nitrogen groups between tissues. Some pyruvate may also be converted to the four-carbon compound oxaloacetate by the incorporation of CO2 in a reaction catalysed by pyruvate carboxylase. This conversion to oxaloacetate can be the first step in the resynthesis of glucose by the process of gluconeogenesis. Alternatively, this may be important as an anaplerotic reaction; these are reactions that maintain the intracellular concentration of crucial intermediates that might otherwise become depleted.

Carbohydrate utilization in different tissues Some tissues, including red blood cells, the renal medulla and the retina, have no mitochondria and therefore no capacity for oxidative metabolism. These tissues therefore rely solely on glycolysis for energy production. Skeletal muscle can derive most of its energy requirement from oxidative metabolism or from anaerobic metabolism, and the choice of fuel depends on the energy demand, the metabolic capacity of the tissue, the availability of substrate and the availability of oxygen. Although conversion of glucose to lactate is an anaerobic process, it occurs even when oxygen is freely available to the muscle, and lactate release does not necessarily imply that the oxygen supply is inadequate. Low forces require recruitment of only some of the type 1 (high aerobic, low glycolytic) muscle fibres, which rely on oxidative metabolism. At higher force requirements, the muscle begins to recruit some of the type II (high glycolytic, lower aerobic) fibres. A point is reached where pyruvate is formed by glycolysis faster than the pyruvate can enter the TCA cycle. The excess pyruvate must be reduced to lactate to allow regeneration of NADþ and continued glycolytic flux. The differing enzyme activities of the different skeletal muscle fibre types offer the opportunity for an exchange of carbohydrate substrate between muscles or between fibres within the same muscle; if the type II fibres are recruited and release lactate into the extracellular space, this might be taken up and oxidized by the adjacent type I fibres, which have a high oxidative capacity. Cardiac muscle has a high oxidative capacity and a low glycolytic capacity, and the heart uses lactate as a fuel for oxidation if this is available.

Regulation of glycolysis The rate of glycolysis must be regulated to ensure that ATP supply is matched to the rate of ATP hydrolysis and with the availability of other energy sources. As well as this local regulation within each cell, there is a need for a coherent response in the different tissues involved in carbohydrate metabolism, and for a coordination of the pathways of carbohydrate metabolism with those of fat and protein. Key steps in the metabolism of carbohydrate include the entry of glucose into the cell, which is regulated by a number of factors. Uptake of glucose from the blood into cells is generally stimulated by insulin, which promotes storage after carbohydrate-containing meals. Transport into muscle is also stimulated by exercise, increasing the availability of glucose as a substrate. Regulation of the rate at which glycolysis proceeds occurs at three key points in the pathway. Hexokinase activity is stimulated by inorganic phosphate, one of the reaction substrates, and is inhibited by the reaction product glucose-6-phosphate. Regulation of phosphorylase is more complex, but it is generally activated by sympathetic activity. PFK is a key regulatory enzyme, and the activity of this enzyme is modified by many different compounds. PFK activity is inhibited by high ATP levels in the cell, and stimulated by increased levels of ADP and adenosine monophosphate (AMP); this means that the activity is low when the cell is energy replete, but high when the energy charge of the cell is low. Citrate can inhibit PFK, allowing a potential link for the integration of fat and carbohydrate

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Gluconeogenesis: the formation of glucose from noncarbohydrate sources Carbohydrate metabolism continues even in the absence of dietary intake, and some resynthesis of carbohydrate can take place using non-carbohydrate sources. Gluconeogenesis e the synthesis of glucose from non-glucose sources e becomes important during starvation by making glucose available to those tissues, including the brain, that cannot use other fuels. Gluconeogenesis also allows recycling of the lactate produced by tissues such as red blood cells.

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A variety of substrates can contribute to gluconeogenesis. Lactate and pyruvate can be used, as well as the glycerol backbone of triglyceride molecules and the carbon skeletons of some amino acids. Gluconeogenesis takes place mostly in the liver and to a lesser extent in the kidney. The reactions of gluconeogenesis involve the reversal of the reactions of glycolysis where this is possible, but the irreversible reactions are bypassed by a different route. In prolonged fasting, glycerol plays a major role in maintaining glucose supply for those tissues for which it is essential; about 20 g of glycerol per day is released as the body’s fat stores are mobilized, and most is converted to glucose. These reactions can only occur when sufficient ATP is available to the cell and when the reduced/oxidized ratio of NADþ permits. The carbon skeletons of most amino acids can contribute to glucose synthesis, and these amino acids are referred to as glucogenic. These amino acids can be metabolized to pyruvate or to intermediates of the TCA cycle, and can then be used for glucose synthesis. Gluconeogenesis allows the large energy store tied up in proteins of labile tissues, such as the gut, to contribute to maintenance of blood glucose during prolonged fasting. Fatty acids cannot be converted to carbohydrate, so only the glycerol component of triglycerides, which is a very small part of the energy stored in adipose cells, is available for gluconeogenesis.

transport in the liver or brain. The actions of glucagon, which is also secreted by the pancreas, are generally antagonistic to those of insulin; the concentration ratio of these two hormones determines their integrated effect on metabolism. The effects of increased glucagon concentration are to stimulate glycogenolysis and gluconeogenesis in the liver, increasing the availability of blood glucose, and to stimulate lipolysis in adipose tissue, increasing the availability of free fatty acids for uptake by the muscle. Adrenaline activates the membrane-bound enzyme adenyl cyclase, increasing the rate of cyclic AMP formation within target cells. An increase in the intracellular cyclic AMP concentration activates phosphorylase and increases the rate of glycogen breakdown in active muscle; stimulation of glycogenolysis in resting muscle is largely prevented because of the low intracellular free calcium levels. The catecholamines also stimulate glycogenolysis in liver and lipolysis in adipose tissue, effectively mobilizing fuels that the muscles can use. Recent research indicates that interleukin-6 and cytokines released from contracting muscle exert a similar effect during exercise. Several other hormones and peptides also play important roles in fuel storage and mobilization. These include growth hormone, cortisol, somatostatin and thyroid hormone. After ingestion of a carbohydrate meal, the insulin concentration rises sharply to promote the disposal of the glucose that appears in the blood, and the glucagon concentration falls. In response to food intake, a large number of small peptides are released by the intestine and some of these stimulate insulin secretion. Anabolic reactions are therefore promoted and catabolic reactions suppressed after feeding. A

Hormonal control of carbohydrate metabolism Several hormones play crucial roles in the regulation of carbohydrate utilization as well as in integrating the supply and utilization of carbohydrate fuels in different tissues. The blood glucose concentration must be maintained within narrow limits, and cannot be allowed to rise too far after a high-carbohydrate meal or to fall too far during fasting or muscular exercise. Insulin is central to the regulation of carbohydrate metabolism. It also has a regulatory function in lipid and protein metabolism, so it is plays a key role in the body’s fuel homoeostasis. Secretion of insulin by the b-cells of the pancreas is stimulated by increasing blood glucose concentrations. In skeletal and cardiac muscle, increasing insulin concentrations increase the number of active glucose transporters and thus stimulate glucose uptake, but insulin has no effect on glucose

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FURTHER READING Barasi ME. Human nutrition. London: Arnold, 2003. Maughan RJ, Gleeson M, Greenhaff PL. Biochemistry of exercise and training. Oxford: Oxford University Press, 1997. Newsholme EA, Leech AR. Biochemistry for the medical sciences. Chichester: Wiley, 1983.

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