From Monosaccharides to Glycogen Synthesis PDF

a wave that consists of oscillating electromagnetic fields radiating outward at the speed of light Carbohydrates The relationship of monosaccharides - polysaccharides is analogous to that of amino acids -proteins, or...

a wave that consists of oscillating electromagnetic fields radiating outward at the speed of light Carbohydrates The relationship of monosaccharides - polysaccharides is analogous to that of amino acids -proteins, or nucleotides - nucleic  Carbohydrates: trasformations of light energy They are mainly produced by photosynthesis They are the most abundant class among the molecules of biological origin They are an essential component for all living organisms They account for more than 90% of the dry 12H 2O + 6CO 2 ⎯energia ⎯ ⎯luminosa ⎯⎯ ⎯→ 6O 2 + C6 H 12O 6 + 6H 2O matter of plants They contribute for the 55 - 70% of the caloric intake in humans 6  The Building Block: Their chemical Chiral tetrahedral carbon behavior depends on atom, covalently linked to a the simultaneous hydroxyl group, a carbonyl presence of a group and the rest of the carbonyl group chain. From a chemical (aldehyde or ketone) point of view they are and (various) aldehydes or ketones of hydroxyl groups: aliphatic polyhydroxy cyclization alcohols with a number of Glyceraldehyde carbon atoms between 3 They possess one or and 7. Each molecule two ends displaying a displays several chiral non-chiral alcohol centers.In nature, D functionality stereoisomerism is prevalent. Glyceraldehyde Glyceraldehyde is the simplest aldose displaying optical activity (C2 asymmetric) The two enantiomers are indicated by D (+) and L (-) Conventionally, the two stereoisomers of glyceraldehyde are used as a reference to classify all monosaccharides CHO CHO H OH HO H CH2OH CH2OH D-glyceraldehyde L-glyceraldehyde 8 Enantiomers Monosaccharides They are aldehydes or ketones of aliphatic polyhydroxy alcohols with a number of carbon atoms between 3 and 7 In all the monosaccharides belonging to the D series , such as in the case of glyceraldehyde, the hydroxyl bound to the asymmetric carbon furthest from the carbonyl group is positioned on the right. 9 Classification of monosaccharides Monosaccharides are classified according to the nature of the carbonyl group (aldehyde or ketone) and the number of carbon atoms a) the trioses b) two hexoses 10  Stereoisomerism Monosaccharides possess numerous chiral centers The number of possible stereoisomers is 2n-2 for aldoses and 2n-3 for ketoses Monosaccharides that differ only in the configuration of a carbon atom are called epimers 11 Epimers 12  Cyclic conformations Alcohols react with carbonyl groups of aldehydes and ketones to form hemiacetals and hemiketals The hydroxyl and carbonyl groups of 5- and 6-carbon monosaccharides spontaneously react intramolecularly to form cyclic hemiacetals and hemiketals 13 Haworth projections A monosaccharide displaying a 6- carbon ring is called pyranose, while a monosaccharide with a 5-carbon ring is called furanose 14 D(+)-glucose It is a reducing sugar with sweetening power of 70% of sucrose At room temperature, a D- glucose solution exhibits a specific optical rotation of + 52.7° Can frequently react with amino acids and be involved in browning reactions (Maillard reaction) It displays a high solubility in water and is present in fruit and honey (as a product of hydrolysis of sucrose) 15 D(-)-fructose It was I initially called levulose, it possesses a sweetening power of 140%, compared to sucrose At room temperature a solution of D-fructose exhibits a specific optical rotation of [α]D20 = - 92.4° It is present in vegetables (onion and chicory), fruit and honey 16  Glucose cyclization Right= below the ring Left = above the ring s The cyclization of a monosaccharide induces asymmetry in the carbonyl carbon leading to the formation of two diastereoisomers called anomers The anomer α possesses the anomeric hydroxyl in the opposite direction, with respect to the ring, to the CH2OH (glucose C6) bound to the chiral center (glucose C5) which determines the configuration D or L. The other anomer is called β. The two anomers, like any other pair of diastereomers, possess different chemical-physical characteristics 17 Mutarotation D-glucose anomers have different specific optical rotation Anomer α: [α]D20 = + 112.2° Anomer β: [α]D20 = + 18.7° At room temperature (25 ° C) a solution of D-glucose has a specific optical rotation of [α]D20 = + 52.7° The two anomers rapidly interconvert and at equilibrium in water the β anomer represents 63.6% of D-glucose while the α anomer 36.4%. The open-chain form of glucose in solution is about 0.02%. 18 The conformation of monosaccharides The pyranose and furanose rings of monosaccharides are not planar since all the carbon atoms possess sp3 hybridization The stability of the two conformations depends on the stereochemical interactions between the substituents of the ring 19  Derivatives of monosaccharides Deoxy-sugars: one or more hydroxyl group is replaced by an H. Amino-sugars: one or more -OH groups are replaced by amino groups (often acetylated) 20  Glucose family Besides oligosaccharides and polysaccharides a monosaccharide can act as “skeleton” for developing novel biological entities. In general, one or more hydroxyls are substituted by other functional groups providing new chemical properties to the overall molecular structure. The relationship of these substituents to monosaccharides is analogous to that of side chains and amino acids. These reactions are important in metabolic processes Leaving Electrophilic site Nucleophilic group attacking agent 21 Remind the nucleophilic acyl substitution  Redox reactions Carbohydrates display the typical reactivity of aldehydes and ketones The mild oxidation of the aldonic carbonyl (C1) group of an aldose generates an aldonic aldaric acid uronic The oxidation of the last alcohol group (C6) gives rise to an uronic acid The oxidation of both the last alcohol group and the Bear in mind that in organic chemistry and in aldehyde group gives rise to an biochemistry oxidation corresponds to a loss of hydrogens or to the gain of oxygens. aldaric acid 22  Lactonization Both aldonic and uronic acids are prone to the intramolecular esterification forming 5 or 6- membered lactones Once again a nucleophilic acyl substitution 23  Ascorbic acid Vitamin C is a γ-lactone of an hexonic acid displaying an enediol structure at carbon atoms 2 and 3 24 Reduction reactions H O H [H] H OH R R The aldoses and ketoses can be reduced under mild conditions to give the corresponding alditols. They are produced by reduction of monosaccharides, by chemical or by enzymatic reactions. They are not monosaccharides because they do not have a carbonyl group. They are commonly found in nature They generate the same caloric intake of monosaccharides but they are absorbed very slowly. Recently, these substances has generated interest, due to their high sweetening power as low calorie sweeteners (GRAS) and their hygroscopicity. 25 D(-)-sorbitol It results from the reduction of glucose It is common in plants In humans, it can be slowly metabolized, providing the same caloric intake of glucose (~ 4 kcal / g) without increasing blood sugar It is not fermentable by yeasts Industrial uses (jams) High hygroscopicity Honey aspect High thermostability Retardant of the crystallization of glucose and sucrose Sweetening power 70% of sucrose Laxative effects 26 Xilitol It is found in small quantities in some fruits and vegetables It looks like sucrose and provides the same sweetening power and the same caloric intake It is not cariogenic In humans it can be incompletely converted into glucose (20 - 80%) and is used in diabetic foods 27  Glycosidic bond The derivatives formed by the reaction between a monosaccharide and an alcohol is called a glycoside The product is stable in alkaline and oxidant conditions, but since the reaction is reversible, the glycosides can be hydrolyzed in an acidic environment In nature, glycosides are very common (they can be distinguished into a glyconic and an aglyconic component) The bond that connects the anomeric carbon with an alcoholic oxygen is defined glycosidic bond This bond is used to obtain polymers (polysaccharides) and represents the analogue of the peptide bond in polypeptide chains 28 Glycosides The derivative formed by the reaction between a monosaccharide and an alcohol is called a glycoside The product is stable towards bases and oxidants, but since the reaction is reversible, glycosides can be hydrolysed in an acid medium In nature, glycosides are very common (a glyconic and an aglyconic component are distinguished) 29  Glycosides Also known as heterosides, pro-drugs: once taken they undergo enzymatic hydrolysis processes which separate the sugar part from the aglycone. The latter generally represents the pharmacologically active fraction of the molecule; the sugar part, however, helps to modulate the intensity of action, its toxicity and the solubility of the entire molecule. 30  Glycosides Amigdalina Amygdalin/Laetrile the most important of the cyanogenic glycosides. It is contained in the seeds of various Rosaceae and, in large quantities, above all in bitter almonds, where it is present in the ratio of 2.5-3.5%. Other parts containing this glycoside are fruits, leaves, flowers and bark of many plants belonging Laetrile to the Rosaceae family. Amygdalin is a cyanogenic glycoside, i.e. capable of releasing hydrogen cyanide Amygdalin/laetrile ("vitamin 17" amygdalin with one less glucose) has been the subject of numerous studies in order to verify its alleged and never demonstrated beneficial effect against cancer. Streptomycin Streptomycin is a bacteriostatic antibiotic at therapeutic doses, Streptomicina at higher doses it becomes bactericidal. It inhibits protein synthesis in ribosomes by binding to RNA Ouabain Ouabain is a poison that binds and inhibits the activity of the Na+/k+ pump 31  Reducing means that a sugar can Reducing sugars give electrons to an oxidant or, in Fehling other the words, it can be oxidized. The term refers to the availability of the carbonyl group, which can be oxidized once the cycle is opened. When the anomeric carbon is involved in a glycosidic bond the cycle can’t be opened, the carbonyl group can’t C-1 is an anomeric carbon be obtained and, as a but it is not oxidizable consequence, it can’t be oxidized. because it is involved into a glycosidic bond In a long polysaccharide (e.g. amylose) the reducing end is the end displaying an anomeric oxidizable carbon (C-1) not involved into This C-1 is the only any glycosidic bond and, for this anomeric carbon not C-4 is not an anomeric carbon involved into a glycosidic reason, oxidizable. bond, hence it is oxidizable  Reducing, non-reducing ends, branches Branch Non-reducing end Reducing end Tollens An alternative of the Fehling reaction 33 Disaccharides They are homo- or heterodimers of monosaccharides Lactose: β-D-galactopyranosyl-(1→4)-β-D-glucopyranose Sucrose: β-D-fructofuranosyl-(1→2)-α-D-glucopyranoside Trehalose: α-D-glucopyranosyl-(1→1)-α-D-glucopyranoside 34  Polysaccharides Homopolysaccharides Hetero-polysaccharides linear branched linear branched 35 The main polysaccharides 36 Reserve polysaccharides: starch Starch represents the glucose reserve in plants and the most common food for humans Glucose is stored in the form of a polymer because the storage of single molecules would create enormous osmotic pressure The polymer chains are packed to avoid an increase in the viscosity of the intracellular medium Starch is made up of a mixture of two polysaccharides Amylose Amylopectin 37 Starch 38 Amylose Amylose consists of a linear chain of several hundred units of glucose linked by α-(1→4) glycosidic bond The average molecular weight is ~ 106 Amylose takes on a helical conformation 39 Amylopectin As amylose, it is constituted of glucose molecules connected by α-(1→4) glycosidic bond. It also presents α-(1→6) ramifications every 24 - 30 glucose units The polymer consists of approximately 106 glucose units with a total molecular weight of approximately 108 40 Starch structure 41  Starch crystal structures Amylose and the outermost polysaccharide chains of amylopectin polymers form helical structures that aggregate in crystalline form These structures are not attacked by hydrolytic enzymes 42 Starch granules 43  Starch granules ilo They display a concentric lamellar structure with alternating crystalline and amorphous phases The crystalline phases are due to the regular packing of the amylopectin terminal chains 44 Starch gelatinization Starch is scarcely hydrated and can not be attacked by hydrolytic enzymes The heating in water causes the disintegration of the crystalline phase of the granules and consequent hydration Once hydrated, amylose and amylopectin can be hydrolyzed 45  Starch retrogradation Upon heating, the starch granules absorb water If the solution is cooled again, it gels due to the formation of inter-chain H bonds between the amylose and amylopectin molecules. Finally, the retrogradation process can be observed 46 Structural polysaccharides: cellulose It represents about half of all organic carbon present on the earth's surface (1015 Kg/year) Linear polymer of about 15,000 D-glucose units connected by β-(1→4) glycosidic bond It does not have a defined size as there is no genetic reference that controls its synthesis It is enzymatically hydrolyzed by cellulases. In many herbivorous animals such as ruminants cellulases are produced by symbiotic bacteria. Endogenous cellulases are produced by a few types of metazoan animals, such as some termites, snails and earthworms 48 Structural polysaccharides: differences between amylose and cellulose The β-(1→4) glycosidic bond of cellulose allows the linear arrangement of the monosaccharide The α-(1→4) glycosidic bond of units and the formation of intra-chain hydrogen amylose induces the polymer bonds provide high stability and strength. Despite to assume a helical its hydrophilicity, it is insoluble in water. deformation The length of the chains does not allow perfect parallelism. The alternate arrangement display crystalline regions (parallel chains linked by H bridges) and amorphous regions (irregularly 49 arranged chains) Chitin It is the main component of the exoskeleton of insects, crustaceans and spiders. It is present in the cell wall of many fungi and algae Linear polymer of N-acetyl-D- glucosamine linked by β-(1→4) glycosidic bond Similarly to cellulose, the numerous intra and inter-chain hydrogen bonds confer high stability and strength. Despite its hydrophilicity, it is insoluble in water 50 Nucleosides and Nucleotides Aglyconic and sugar moieties  Nucleosides Nucleosides are glycosylamines made up of two components: 5 carbon atom sugar ring furanose, -amine ribose or 2-deoxyribose) and a Nitrogenous base: nitrogenous base also known as Providing the identity nucleobase. Glycosyl- Ribose ring: Nitrogenous base are five: Purines the carrier, the Nucleoside (two rings), Pyrimidines (one ring). spacer. «Glycosyl-amine» The backbone unit  Nucleotides Nucleotides are the building blocks of DNA and RNA. They are made up of three components: a five- carbon sugar, a nitrogenous base, and one or more phosphate groups. Nucleoside + phosphate group= Nucleotide Living organisms naturally encode heritable information using just four bases of DNA: adenine (A), thymine (T), cytosine (C) and guanine (G). In RNA , thymine is replaced with uracil. Organisms use messenger RNA (mRNA) to convey genetic information using guanine (G), uracil (U), adenine (A), and cytosine (C)that directs synthesis of specific proteins. Many viruses encode their genetic information using an RNA genome. A universal function in which RNA molecules direct the synthesis of proteins on ribosomes. This process uses transfer RNA (tRNA) molecules to deliver amino acids to the ribosome, where ribosomal RNA (rRNA) then links amino acids together to form coded proteins. Nucleotides: DNA and RNA Phosphodiester bond building blocks A nucloetide is the repeating unit of the DNA or RNA polymer Ribonucleotides are in RNA Deoxyribonucleotides are in DNA The nitrogen base is attached β to Ribose (RNA) Deoxyribose (DNA) In the DNA structure this hydroxyl is bound with the The sugar is phosphorylated at carbon 5' phosphate belonging to another nucleotide Phosphodiester bond Phosphoester bond Phosphodiester bond Nucleophilic substitution: The nucleophile is the 3' oxygen at the 3' end of the growing DNA strand. Electrons from this oxygen are used to form a new bond with the phosphorus atom of the α-phosphate group of the incoming nucleotide. Complementary base pairing  DNA structure The relationship of bases to nucleotides is analogous to that of side chains and amino acids. Adenine, Cytosine, Guanine and Tyrosine, in DNA determines our unique genetic code and provides the instructions for producing molecules in the body. The cell reads the DNA code in groups of three bases Complementary base pairing Phosphodiester bond: the backbone Nucleotides and energy: phosphoanhydride bonds In addition to be building blocks for the construction of nucleic acid polymers, singular nucleotides play roles in cellular energy storage and provision. They provide chemical energy—in the form of the nucleoside triphosphates, adenosine triphosphate (ATP), guanosine triphosphate (GTP), cytidine triphosphate (CTP) and uridine triphosphate (UTP). The phoshodiester bond is the junction of the DNA structure. In ATP, UTP etc. for energy purposes: phosphoanhydride bonds Adenin Reaction coupling kJ/mol kJ/mol In the first reaction, a phosphate group is transferred from ATP to glucose, forming a phosphorylated glucose intermediate (glucose-P). This is an energetically favorable (energy-releasing) reaction because ATP is so unstable, i.e., really "wants" to lose its phosphate group. In the second reaction, the glucose-P intermediate reacts with fructose to form sucrose. Because glucose-P is relatively unstable (thanks to its attached phosphate group), this reaction also releases energy and is spontaneous.  Glycogen and glucose regulation The storage form of carbohydrates in animals is known as glycogen It is most abundant in muscle fibers and the liver Similar in structure to amylopectin but more extensively branched (one α- (1→6) bond for every 8-12 residues) Intracellularly degraded by glycogen phosphorylase to yield glucose-1- phosphate Two hormones control the Blood glucose concentration of glucose in the regulation blood Insulin: stimulates glycogen synthesis, lipogenesis and glycolysis Glucagon:stimulates glycogen degradation and lipolysis 62 Glycogen Synthase vs Glycogen Phosphorylase  Glucose transformations before glycogen synthesis: Each step requires a specific enzyme 1) The main reason for the immediate phosphorylation of glucose is to prevent diffusion out of the cell. The phosphorylation adds a charged phosphate group so the glucose 6-phosphate cannot easily cross the cell membrane. 2) C1 must bear the phosphate because it must be activated for creating the glycosidic bond. Phosphoglucomutase Substrate recognition: induced fit The enzyme changes shape following the interaction with the substrates to form an Enzyme-substrate complex. the binding of Hexokinase with its substrates (ATP and sugar) is accompanied by a significant conformational modification Glycogen synthesis strategy This carbon is not reactive. How is it possible to break the bond with the OH group? Conversion of glucose-1-phosphate into UDP-glucose Reactive! The instability of UDP-Glc is actually related to the presence of the phosphoanhydride bond. The system does not allow UDP-Glc to be stabized through its hydrolysis. Thus, the only way of lowering the overall energy is the sobstituion of UDP with the glycosidic bond of the 68 elongating glycogen chain UDP  Glycogen synthesis The nucleophilic sobstitution C1 C4 reaction does not take place is The synthesis of glycogen is mediated by the enzyme nucleophile carried out by glycogen synthase which uses UDP-glucose as the first substrate (anomeric C1 electrophilic site ) and the non- reducing end of glycogen as the second (C4). The first glucose residue is provided by a protein, called glycogenin, which self-glycosiles on a tyrosine residue and which remains localized in the center of the glycogen molecule. 70 Glycogen «de novo» synthesis Glycogen «de novo» synthesis Glycogen synthase mechanism Regulation of the glycogen synthesis Glycogen synthase is a homotetrameric enzyme. The enzymatic activity is controlled by phosphorylation of a serine residue. This phosphorylation reduces glycogen synthase activity in the phosphorylated state (glycogen synthase b) the enzyme requires the allosteric activator glucose-6-phosphate. 75 Regulation of glycogen hydrolysis The degradation of glycogen is catalyzed by glycogen phosphorylase (a 97 kDa homodimer that has pyridoxal phosphate as a cofactor) The enzyme uses inorganic phosphate to give glucose-1- phosphate The activity of glycogen phosphorylase is also modulated by phosphorylation 76 77  Glycogen phosphorylase The mechanism involves the formation of a ternary complex (E-phosphate-glycogen) with the consequent formation of a carbocationic intermediate The release of glucose-1-phosphate follows Vitamin B6 the coenzyme the reaction between the intermediate and the phosphate present in the active site 78  Glycogen synthase and phosphorylase are also under the control of allosteric effectors: AMP, ATP, Glucose-6 phosphate Glycogen synthase is activated by ATP and Glycogen synthase is activated by AMP Glucose-6 phosphate Glycogen synthase INHIBITED Glycogen synthase ACTIVATED Glycogen phosphorylase ACTIVATED Glycogen phosphorylase INHIBITED HIGH ENERGY ACTIVATES GLYCOGEN LOW ENERGY ACTIVATES GLYCOGEN SYNTHESIS (Glucose/energy storage) HYDROLISIS  Starch sinthesys Sucrose is the end product of photosynthesis Inside the plant cell it is hydrolyzed to glucose and fructose (the latter is converted into glucose by an isomerase) Glucose is transformed into uridine diphosphate glucose (udpg) Then it is transformed into glucose-1- phosphate (G-1-P) Phosphoglucomutase transforms it into glucose-6-phosphate which can be translocated within the amyloplast In the amyloplast it is re-transformed into G-1-P and finally into glucose-adenosine- diphosphate from which it is possible to synthesize amylose and amylopectin 80