Lehninger. Stryer. Γεωργάτσος. Internet

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1 Lehninger Stryer Γεωργάτσος Internet

2 1800 Πέψη κρέατος και αμύλου από βιολογικά υγρά στομάχου 1850 Louis Pasteur Ζύμωση σακχάρων, μήκυτες απαραίτητοι 1897 Eduard Buchner υδρόλυμα μηκύτων Nobel James Sumner καθαρισμός ουρεάσης 1930 John Northop & Stanley κρυστάλλωσαν pepsin & trypsin Nobel 1946 Σήμερα πάνω από 2000 ένζυμα καλά χαρακτηρισμένα

3 Κρυσταλλογράφος Χάρτης αντανάκλασης Ηλεκτρονική πυκνότητα Τρισδιάστατη δομή πρωτεϊνών

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7 ΕΝΖΥΜΑ ΚΑΤΑΤΑΞΗ ΚΑΙ ΟΝΟΜΑΣΙΑ ΤΩΝ ΕΝΖΥΜΩΝ 1. Οξειδορεδουκτάσες 2. Τρανσφεράσες 3. Υδρολάσες 4. Λυάσες 5. Ισομεράσες 6. Λιγάσες Φωσφουδρολάση ορθοφωσφορικών μονοεστέρων Αλκαλική φωσφατάση (κοινή ονομασία) =υδρολάσες 1=υδρολάσες που δρούν σε εστερικούς δεσμούς 3= υποομάδα που δρα σε φωσφορικούς μονοεστέρες 1= ειδικό ένζυμο υποομάδας = όξινη φωσφατάση

8 EC 1 Oxidoreductases EC 1.1 Acting on the CH-OH group of donors EC1.2 Acting on the aldehyde or oxo group of donors EC 1.3 Acting on the CH-CH group of donors EC 1.4 Acting on the CH-NH 2 group of donors EC 1.5 Acting on the CH-NH group of donors EC 1.6 Acting on NADH or NADPH EC 1.7 Acting on other nitrogenous compounds as donors EC 1.8 Acting on a sulfur group of donors EC 1.9 Acting on a heme group of donors EC 1.10 Acting on diphenols and related substances as donors EC 1.11 Acting on a peroxide as acceptor EC 1.12 Acting on hydrogen as donor EC 1.13 Acting on single donors with incorporation of molecular oxygen (oxygenases) EC 1.14 Acting on paired donors, with incorporation or reduction of molecular oxygen EC 1.15 Acting on superoxide radicals as acceptor EC 1.16 Oxidising metal ions EC 1.17 Acting on CH or CH 2 groups EC 1.18 Acting on iron-sulfur proteins as donors EC 1.19 Acting on reduced flavodoxin as donor EC 1.20 Acting on phosphorus or arsenic in donors EC 1.21 Acting on X-H and Y-H to form an X-Y bond EC 1.97 Other oxidoreductases

9 EC 2 Transferases EC 2.1Transferring one-carbon groups EC 2.2Transferring aldehyde or ketonic groups EC 2.3Acyltransferases EC 2.4Glycosyltransferases EC 2.5Transferring alkyl or aryl groups, other than methyl groups EC 2.6Transferring nitrogenous groups EC 2.7Transferring phosphorus-containing groups EC 2.8Transferring sulfur-containing groups EC 2.9Transferring selenium-containing groups EC 3 Hydrolases EC 3.1 Acting on ester bonds EC 3.2Glycosylases EC 3.3Acting on ether bonds EC 3.4Acting on peptide bonds (peptidases) EC 3.5Acting on carbon-nitrogen bonds, other than peptide bonds EC 3.6Acting on acid anhydrides EC 3.7Acting on carbon-carbon bonds EC 3.8Acting on halide bonds EC 3.9Acting on phosphorus-nitrogen bonds EC 3.10Acting on sulfur-nitrogen bonds EC 3.11Acting on carbon-phosphorus bonds EC 3.12Acting on sulfur-sulfur bonds EC 3.13Acting on carbon-sulfur bonds

10 EC 4 Lyases EC 4.1Carbon-carbon lyases EC 4.2Carbon-oxygen lyases EC 4.3 Carbon-nitrogen lyases EC 4.4 Carbon-sulfur lyases EC 4.5 Carbon-halide lyases EC 4.6 Phosphorus-oxygen lyases EC 4.99 Other lyases EC 5 Isomerases EC 5.1 Racemases and epimerases EC 5.2 cis-trans-isomerases EC 5.3 Intramolecular isomerases EC 5.4 Intramolecular transferases (mutases) EC 5.5 Intramolecular lyases EC 5.99 Other isomerases EC 6 Ligases EC 6.1 Forming carbon oxygen bonds EC 6.2 Forming carbon sulfur bonds EC 6.3 Forming carbon nitrogen bonds EC 6.4 Forming carbon carbon bonds EC 6.5 Forming phosphoric ester bonds EC 6.6 Forming nitrogen metal bonds

11 Οξειδορεδουκτάσες Oxidoreductase is an enzyme that catalyzes the transfer of electrons from one molecule (the reductant, also called the hydrogen acceptor or electron donor) to another (the oxidant, also called the hydrogen donor or electron acceptor). For example A + B A + B In this example, A is the reductant (electron donor) and B is the oxidant (electron acceptor). δευδρογονάσες καταλύουν την μεταφορά υδρογόνου, δότες υδρογόνου CHOH, CHO, CHO-CH, CH-NH2, CH-NH κλπ δέκτες NAD, NADP οξειδάσες εάν ο αποδέκτης είναι μοριακό οξυγόνο υπεροξειδάσες χρησιμοποιούν το Η 2 Ο 2 ως οξειδωτικό παράγοντα υδροξυλάσες στο υπόστρωμα τους άτομα οξυγόνου από μοριακό οξυγόνο οξυγονάσες ενσωματώνουν ολόκληρο μόριο οξυγόνου σε διπλούς δεσμούς Τρανσφεράσες Τransferase is an enzyme that catalyzes the transfer of a functional group (e.g. a methyl or phosphate group) from one molecule (called the donor) to another (called the acceptor). For example, an enzyme that catalyzed this reaction would be a transferase: A X + B A + B X In this example, A would be the donor, and B would be the acceptor. The donor is often a coenzyme Ένζυμα που μεταφέρουν μονοανθρακικές ομάδες, ακυλομάδες, γλυκοσυλομάδες, αζωτούχες, φωσφορικές, θειούχες κλπ

12 Υδρολάσες Hydrolase is an enzyme that catalyzes the hydrolysis of a chemical bond. For example, an enzyme that catalyzed the following reaction is a hydrolase: A B + H2O A OH + B H Διασπούν μεγάλη ποικιλία δεσμών εστεράσες, φωσφατάσες, γλυκοσιδάσες, πεπτιδάσες Λυάσες lyase is an enzyme that catalyzes the breaking of various chemical bonds by means other than hydrolysis and oxidation, often forming a new double bond or a new ring structure. For example, an enzyme that catalyzed this reaction would be a lyase: ATP camp+ PPi αλδολάσες που διασπούν τον δεσμό C-C, αποκαρβοξυλάσες, κετοξυλυάσες Ισομεράσες isomerase is an enzyme that catalyses the interconversion of polymers. Isomerases thus catalyze reactions of the form A B Ρακεμάσες, επιμεράσες, cis-trans ισομεράσες πχ ισομεράση της γλυκόζης Λιγάσες ligase (from the Latin verb ligāre "to bind" or "to glue together") is an enzyme that can catalyse the joining of two large molecules by forming a new chemical bond, usually with accompanying hydrolysis of a small chemical group pendant to one of the larger molecules. Generally ligase catalyses the following reaction: Ab + C A C + b or sometimes Ab + cd A D + b + c where the lower case letters signify the small, pendant groups. Δημιουργία δεσμών μεταξύ δύο μορίων με δότη ενέργειας ATP

13 Κυτοχρώματα: πρωτείνες που περιέχουν αίμη, ρόλος μεταφορά ηλεκτρονίων, τα πιο γνωστά a 1, a 3, b, c, cc 1 Κυτόχρωμα c 12400Da, 104 αμινοξέα

14 Δύο δεσμοί κυστεινών με τους πυρολικούς δακτυλίους της αίμης Ι και ΙΙ

15 Αποένζυμο, ολοένζυμο, συνένζυμα συνδέονται στοιχειομετρικά με το αποένζυμο και το υπόστρωμα. Οι βιταμίνες εξωγενείς διαιτητικοί παράγοντες Συνένζυμα οξειδορεδουκτασών: νουκλεοτίδια του νικοτιναμιδίου NAD+ NADP Βιταμίνη νικοτιναμίδιο ΔΗ 2 + NAD + Δ+NADH+ H + όπου ΔΗ2 δότης υδρογόνου, κύριος ρόλος η μεταφορά υδρογόνου στην αναπνευστική αλυσίδα Nicotinamide adenine dinucleotide (NAD + ) is an important coenzyme found in cells. It plays key biochemical roles as a carrier of electrons and a participant in metabolic redox reactions, as well as in cel signaling. There are two forms of this coenzyme in cells, NAD + and the phosphorylated form NADP +. These two related coenzymes have similar chemistry, but perform different roles in metabolism.

16 Φλαβινοσυνένζυμα = ριβοφλαβίνη, η αναγωγή των συνενζύμων λαμβάνει χώρα με την προσθήκη υδρογόνου στα άτομα Ν * Riboflavin (E101), also known as vitamin B2, is an easily absorbed micronutrient with a key role in maintaining health in animals. It is the central component of the cofactors FAD and FMN, and is therefore required by all flavoproteins. As such, vitamin B2 is required for a wide variety of cellular processes. Like the other B vitamins it plays a key role in energy metabolism, and is required for the metabolism of fats, carbohydrates, and proteins. Milk, cheese, leafy green vegetables, liver, legumes such as mature soybeans, yeast and almonds are good sources of vitamin B2, but exposure to light destroys riboflavin.

17 Άλλα οξειδοαναγωγικά συνένζυμα Αίμη, ουβικινόνη This vitamin-like substance is, by nature, present in all human cells and responsible for the production of the body s own energy. In each human cell, food energy is converted into energy in the mitochondria with the aid of CoQ10. Ninety-five percent of all the human body s energy requirements (ATP) is converted with the aid of CoQ10. Therefore, those organs with the highest energy requirements such as the heart, the lungs, and the liver have the highest CoQ10 concentrations. A heme or haem is a prosthetic group that consists of an iron atom contained in the center of a large heterocyclic organic ring called a porphyrin. Not all porphyrins contain iron, but a substantial fraction of porphyrin-containing metalloproteins have heme as their prosthetic subunit; these are known as hemoproteins

18 It is essential for aerobic life and a common dietary supplement. Dihydrolipoic acid, the reduced form of lipoic acid, although it is sometimes also called "lipoic acid." One of the most visible roles of lipoic acid is as a cofactor in aerobic metabolism, specifically the pyruvate dehydrogenase complex. Lipoate participates in transfer of acyl or methylamine groups

19 Γλουταθειόνη Glutathione (GSH) is a tripeptide. It contains an unusual peptide linkage between the amine group of cysteine and the carboxyl group of the glutamate side chain. Glutathione, an antioxidant, protects cells from toxins such as free radicals. Thiol groups are kept in a reduced state within ~5 mm in animal cells. In effect, glutathione reduces any disulfide bonds formed within cytoplasmic proteins to cysteines by acting as an electron donor. Glutathione is found almost exclusively in its reduced form, since the enzyme which reverts it from its oxidized form (GSSG), glutathione reductase, is constitutively active and inducible upon oxidative stress Ασκορβικό οξύ The L-enantiomer of ascorbic acid is also known as vitamin C. The name "ascorbic" comes from its property of preventing and curing scurvy. Primates, including humans, and a few other species in all divisions of the animal kingdom, notably the guinea pig have lost the ability to synthesize ascorbic acid and must obtain it in their food.

20 Συνένζυμα τρανσφερασών: 1. Αδενοσινοτριφωσφορικό οξύ-atp Adenosine 5'-triphosphate (ATP) is a multifunctional nucleotide that is most important as a "molecular currency" of intracellular energy transfer. In this role, ATP transports chemical energy within cells for metabolism. It is produced as an energy source during the processes of photosynthesis and cellular respiration and consumed by many enzymes and a multitude of cellular processes including biosynthetic reactions, motility and cell division. In signal transduction pathways, ATP is used as a substrate by kinases that phosphorylate proteins and lipids, as well as by adenylate cyclase, which uses ATP to produce the second messenger molecule cyclic AMP

21 Φωσφορυλιώνει αλκόολες R-OH + ATP R-O-P + ADP Φωσφορυλιώνει την γουανιδική ομάδα R-NH-C(NH)-NH 2 + ATP R-NH-C(NH)-NH-P + ATP Φωσφορυλιώνει οργανικά οξέα R-COOH + ATP R-CO-P + ADP Πυροφωσφορυλιώνει αλκόολες R-OH + ATP R-O-P-P +AMP Φωσφορυλιώνει νουκλεοτίδια CMP + ATP CDP + ADP Αδενυλιώνει οργανικά οξέα R-COOH + ATP R-CO-AMP + P-P Αδενυλιώνει πρωτείνες Protein-OH + ATP Protein-AMP + P-P Μεταφέρει την αδενοσίνη R-S(CH 3 ) + ATP R-S(CH 3 )-Adenosine + P-P + Pi Ο πιο σημαντικός ρόλος του η φωσφορυλίωση υποστρωμάτων κινασών

22 2. Συνένζυμο Α ή CoA ή CoA-SH μεταφέρει ομάδες οργανικών οξέων. Αποτελείται από 3-5 διφωσφορική αδενοσίνη και φωσφορική παντοθείνη. Η σπουδαιότερη ένωση του το ακέτυλο-συνένζυμο Α Acetyl-CoA is an important molecule in metabolism, used in many biochemical reactions. Its main use is to convey the carbon atoms within the acetyl group to the Krebs Cycle to be oxidized for energy production.

23 3. Ενεργό θειικό= δότης θειικών ομάδων εστεροποίηση φαινολών στεροειδών κλπ

24 4. Ενεργό μεθύλιο διαθέτει την μεθυλοομάδα σε άτομα με ελεύθερα ηλεκτρονιακάζεύγη

25 5. Ενεργό φορμύλιο ή φολικό οξύ μεταφορέας μονοανθρακικών ομάδων όπως -CH 3,-CH 2 OH, -CHO στις θέσεις N 5 και N 10

26 6. Ενεργό καρβοξύλιο ή βιοτίνη δεσμεύει CO 2 και το αποδίδει σε αντιδράσεις καρβοξυλίωσης

27 7. Πυροφωσφορική θεαμίνη ή βιταμίνη Β 1 ή θειαμίνη δρα ως συνένζυμο σε αντιδράσεις μεταφοράς αλδευδο παραγώγων

28 8. Βιταμίνη πυριδοξάλη συμμετέχει στην μεταφορά αμινομάδων σε τρανσαμινώσεις, σε ρακεμιώσεις και σε αποκαρβοξυλιώσεις αμινοξέων Συνένζυμα Υδρολασών, Ισομερασών, Λυασών, Λιγασών

29 Κινητική ενζυμικών αντιδράσεων E + S ES ES E + P k 1 E + S ES E + P k 2 k 3

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44 Industrial applications Enzymes are used in the chemical industry and other industrial applications when extremely specific catalysts are required. However, enzymes in general are limited in the number of reactions they have evolved to catalyse and also by their lack of stability in organic solvents and at high temperatures. Consequently, protein engineering is an active area of research and involves attempts to create new enzymes with novel properties, either through rational design or in vitro evolution

45 Baking industry Brewing industry Fungal alpha-amylase enzymes are normally inactivated at about 50 degrees Celsius, but are destroyed during the baking process. Catalyze breakdown of starch in the flour to sugar. Yeast action on sugar produces carbon dioxide. Used in production of white bread, buns, and rolls. Proteases Biscuit manufacturers use them to lower the protein level of flour. Enzymes from barley are released during the mashing stage of beer production.they degrade starch and proteins to produce simple sugar, amino acids and peptides that are used by yeast for fermentation Industrially produced barley enzymes Widely used in the brewing process to substitute for the natural enzymes found in barley. Amylase, glucanases, proteases: Split polysaccharides and proteins in the malt. Betaglucanases and arabinoxylanases: Improve the wort and beer filtration characteristics. Amyloglucosidase and pullulanases: Low-calorie beer and adjustment of fermentability. Proteases: Remove cloudiness produced during storage of beers. Acetolactatedecarboxylase (ALDC): avoid the formation of diacetyl

46 Dairy industry Starch industry Rennin derived from the stomachs of young ruminant animals manufacture of cheese, used to hydrolyze protein Microbially produced enzyme increasing use in the dairy industry. Lipases is implemented during the production of Roquefort cheese to enhance the ripening of the blue-mould cheese. Lactases Break down lactose to glucose and galactose Papain To soften meat for cooking Glucose Fructose Amylases, amyloglucosideases and glucoamylases: Converts starch into glucose and various syrups Glucose isomerase: Converts glucose into fructose in production of high fructose syrups from starchy materials. These syrups have enhanced sweetening properties and lower calorific values than sucrose for the same level of sweetness Paper industry Amylases, Xylanases, Cellulases and ligninases Degrade starch to lower viscosity, aiding sizing and coating paper. Xylanases reduce bleach required for decolorising; cellulases smooth fibers, enhance water drainage, and promote ink removal; lipases reduce pitch and lignindegrading enzymes remove lignin to soften paper.

47 Biofuel industry Cellulases: Used to break down cellulose into sugars that can be fermented Detergents Proteases: produced in an extracellular form from bacteria Used for presoak conditions and direct liquid applications helping with removal of protein stains from clothes Amylases: Detergents for machine dish washing to remove resistant starch residues. Lipases: Used to assist in the removal of fatty and oily stains Cellulases: Used in biological fabric conditioners Molecular biology Restriction enzymes, DNA ligase and polymerases Used to manipulate DNA in genetic engineering, important in pharmacology, agriculture and medicine. Essential for restriction digestion and the polymerase chain reaction. Molecular biology is also important in forensic science.

48 Σύνδεση με ασθένειες One example is the most common type of phenylketonuria. A mutation of a single amino acid in the enzyme phenylalanine hydroxylase, which catalyzes the first step in the degradation of phenylalanine, results in build-up of phenylalanine and related products. This can lead to mental retardation if the disease is untreated Phenylalanine hydroxylase