MBMB 1224 Lecture 4: Enzymes Intro PDF

LECTURE 4: INTRODUCTION TO ENZYMES Historical perspectives Nomenclature Classification Properties Review of biological functional groups 2 Review of amino acids and protein structure Amino Acids with Aliphatic Side Chains (...

LECTURE 4: INTRODUCTION TO ENZYMES Historical perspectives Nomenclature Classification Properties Review of biological functional groups 2 Review of amino acids and protein structure Amino Acids with Aliphatic Side Chains (non-polar) Amino Acids with Hydroxyl-Containing Side Chains 4 Amino Acids with Aromatic Side Chains 5 Amino Acids with Sulfur-Containing Side Chains 6 Amino Acids with “Basic” Side Chains (Structures are those that predominate at pH 7) 7 Amino Acids with Acidic Side Chains and their Amides (Structures are those that predominate at pH 7) 8 Imino acid 9 REMEMBER: Their names Their structure Their three letter code Their one letter code Protein structural organisation  Enzyme structure Enzymes are proteins They have a globular shape A complex 3-D structure http://www.chem.ubc.ca/personnel/faculty/withers/group/group/begum/Hu man%2520Pancreati%2520Amylase% Human pancreatic amylase  The active site The shape and the chemical environment inside the active site permits a chemical reaction to proceed more easily. © 2017 Paul Billiet ODWS  Introduction to enzymes (1). Much of the early history of biochemistry is the history of enzyme research (2). Biological catalysts were first recognized in studying animal food digestion and sugar fermentation with yeast (brewing and wine making) (3). Ferments (i.e., enzymes, meaning in “in yeast”) were thought (wrongly) to be inseparable from living yeast cells for quite some time (Louis Pasteur) 14 (4). Yeast extracts were found to be able to ferment sugar to alcohol (Eduard Buchner, 1897, who won the Nobel Prize in Chemistry in 1907 for this discovery) (5). Enzymes were found to be proteins (1920s to 1930s, James Sumner on urease and catalase ,“all enzymes are proteins”, John Northrop on pepsin and trypsin, both shared the 1946 Nobel Prize in Chemistry) (6). Catalytic RNA (also called ribozyme ---from ribonucleic acid enzyme, or RNA enzyme) were found in the 1980s (Thomas Cech, Nobel Prize in Chemistry in 1989) 15  Characteristics of Enzymes i. Almost all enzymes are proteins. Enzymes follow the physical and chemical reactions of proteins. ii. They are heat labile. iii. They are water-soluble. iv. They can be precipitated by protein precipitating reagents (ammonium sulfate or trichloroacetic acid). v. They contain 16% weight as nitrogen. 16  Naming Enzymes The name of an enzyme in many cases end in –ase For example, sucrase catalyzes the hydrolysis of sucrose The name describes the function of the enzyme For example, oxidases catalyze oxidation reactions Sometimes common names are used, particularly for the digestion enzymes such as pepsin and trypsin Some names describe both the substrate and the function. For example, alcohol dehydrogenase oxidizes ethanol Enzymes Are Classified into six functional Classes (EC number Classification) by the International Union of Biochemists (I.U.B.). on the Basis of the Types of Reactions That They Catalyze EC 1. Oxidoreductases EC 2. Transferases EC 3. Hydrolases EC 4. Lyases EC 5. Isomerases EC 6. Ligases 18  Principle of the international classification Each enzyme has classification number consisting of four digits: Example, EC: (2.7.1.1) HEXOKINASE 19 EC: (2.7.1.1) these components indicate the following groups of enzymes: 2. IS CLASS (TRANSFERASE) 7. IS SUBCLASS (TRANSFER OF PHOSPHATE) 1. IS SUB-SUB CLASS (ALCOHOL IS PHOSPHATE ACCEPTOR) 1. SPECIFIC NAME ATP,D-HEXOSE-6-PHOSPHOTRANSFERASE (Hexokinase) 20  6 CH 6 CH O PO 2  2O H 2 3 ATP ADP 5 5 O H O H H H H H 4 1 4 1 OH H OH H 2+ Mg OH OH OH OH 3 2 3 2 H OH H e x o k in a s e H OH g lu c o s e g lu c o s e - 6 - p h o s p h a te Hexokinase catalyzes: Glucose + ATP  glucose-6-P + ADP  EC 1. Oxidoreductases Biochemical Activity: – Catalyse Oxidation/Reduction Reactions Act on many chemical groupings to add or remove hydrogen atoms Examples: – Lactate dehydrogenase – Glucose Oxidase – Peroxidase – Catalase – Phenylalanine hydroxylase 22  1. Oxidoreductases Catalyze oxidation-reduction reactions - oxidases - peroxidases - dehydrogenases  EC 2. Transferases Biochemical Activity: – Transfer a functional groups (e.g. methyl or phosphate) between donor and acceptor molecules Examples: – Transaminases (ALT & AST) – Phosphotransferases (Kinases) – Transmethylases – Transpeptidases – Transacylases 24  2. Transferases Catalyze group transfer reactions  EC 3. Hydrolases Biochemical Activity: – Catalyse the hydrolysis of various bonds Add water across a bond Examples: – Protein hydrolyzing enzymes (Peptidases) – Carbohydrases (Amylase, Maltase, Lactase) – Lipid hydrolyzing enzymes (Lipase) – Deaminases – Phosphatases 26  3. Hydrolases Catalyze hydrolysis reactions where water is the acceptor of the transferred group - esterases - peptidases - glycosidases  EC 4. Lyases Biochemical Activity: – Cleave various bonds by means other than hydrolysis and oxidation – Add Water, Ammonia or Carbon dioxide across double bonds, or remove these elements to produce double bonds Examples: – Fumarase – Carbonic anhydrase 28 4. Lyases  EC 5. Isomerases Biochemical Activity: – Catalyse isomerization changes within a single molecule – Carry out many kinds of isomerization: L to D isomerizations Mutase reactions (Shifts of chemical groups) Examples: – Isomerase – Mutase 30  5. Isomerases Catalyze isomerization reactions  EC 6. Ligases Biochemical Activity: – Join two molecules with covalent bonds Catalyse reactions in which two chemical groups are joined (or ligated) with the use of energy from ATP Subclasses and the bonds formed by ligases 6.1 C-O 6.2 C-S 6.3 C-C 6.4 C-N 6.5 phosphate ester Examples: – Acetyl~CoA Carboxylase – Glutamine synthetase  6. Ligases (synthetases) Catalyze ligation, or joining of two substrates Require chemical energy (e.g. ATP) Synthetases are ATP-dependent enzymes catalysing biosynthetic reactions; they belong to Ligases (class 6).Examples are Carbamoyl phosphate synthetase; Argininosuccinate synthetase; PRPP synthetase and Glutamine synthetase Synthases are enzymes catalysing biosynthetic reactions; but they do not require ATP directly; they belong to classes other than Ligases. Examples are Glycogen synthase and ALA synthase 34  Properties of enzymes Catalytic efficiency – high efficiency, 103 to 1017 faster than the corresponding uncatalyzed reactions Specificity - high specificity, interacting with one or a few specific substrates and catalyzing only one type of chemical reaction Mild reaction conditions- 37 ℃, physiological pH, ambient atmospheric pressure 35  High efficiency For carbonic anhydrase, an enzyme catalyzing the hydration of CO2 (H2O + CO2 HCO3- + H+), the rate enhancement is 107 (each enzyme molecule can hydrate 105 molecules of CO2 per second!)  High specificity 1). Absolute specificity: the enzyme will catalyze only one reaction. NH 2 H 2O e.g C O 脲酶 Urease CO 2 + 2NH3 NH 2 NH 2 H 2O C O X 脲酶 Urease NHCH 3 2). Relative specificity (i) Group specificity: the enzyme will act only on molecules that have specific functional groups, such as amino, phosphate and methyl groups. A—B or A—B e.g α-D-glucosidase CH 2 OH O OH O R OH OH 2). Relative specificity… ii). Bond specificity: the enzyme will act on a particular type of chemical bond regardless of the rest of the molecular structure. O esterase 酯酶 R 1C + H 2O R 1C O O H + R 2O H OR 2  3). Stereospecificity: The enzyme will act on a particular steric or optical isomer H C COOH fumarate hydratase 延胡索酸水化酶 CH 2 COOH + H 2O HOOC C H CHOHCOOH malate Next lecture Enzyme purification 41