Lecture 8-Enzymes PDF

 11/15/2023 Lecture 8 Taibah University Enzymes Dr. Mekky Abouzied...

 11/15/2023 Lecture 8 Taibah University Enzymes Dr. Mekky Abouzied Dr. Heba Eltahir Objectives  After studying this lecture, the students should be able to: 1. Identify the characteristics of enzyme catalysts. 2. Undrastand the properties of enzymes. 3. Explain how the enzymes accelerate the rate of reactions. 4. Undersatand the enzyme composition and nomenclature. 5. Identify the IUB classification of enzymes. 6. Know the factors affecting enzymes activities. 7. Understand the rule of clinical enzymology in diagnosis of diseases. 1  11/15/2023 Importance Location within the cell Enzymes play an important role in Many enzymes are Metabolism, Diagnosis, and localized in specific Therapeutics. organelles within the cell (see figure). Metabolism Such compartmentalization All biochemical reactions are enzyme serves to isolate the catalyzed in the living organism. reaction substrate or Diagnosis product from other Levels of enzymes in blood are of competing reactions. diagnostic importance e.g. it is a good indicator in diseases such as This provides a favorable myocardial infarction. environment for the Therapeutics. reaction, and organizes the Enzyme can be used therapeutically thousands of enzymes such as digestive enzymes or present in the cell into commercially as in detergents. purposeful pathways. Important Definitions Catalyst: They are organic or inorganic substance that accelerate the rate of chemical reactions Enzyme:  Biological catalysts which speed up the rate of reaction without becoming part of the reaction  Most enzymes are protein in nature that are produced by living cells to serve as biological catalysts.  They catalyze nearly all the chemical reactions taking place in the cells of the body. 2  11/15/2023 Active site  The area on the enzyme where the substrates attach to is called the active site.  Enzymes are usually very large proteins Enzyme molecules contain a special pocket or cleft called the active sites which represent a small region of the enzyme molecule. General characteristics of enzymes 1- Enzymes are neither consumed nor produced during the course of a reaction. 2- Enzymes are mostly proteins. Ribozymes …….??? 3- enzymes are specific enzymes have varying degrees of specificity for substrates Enzymes may recognize and catalyze: - a single substrate - a group of similar substrates - a particular type of bond 4- Enzymes function within a moderate pH and temperature range. 3  11/15/2023 Enzyme's compositions  Enzymes are either simple or conjugated proteins. 1. Simple enzyme: its hydrolysis gives only amino acids. Only native conformation of protein is required for the activity of this type. 2. Conjugated enzyme (holoenzyme): This type composed of protein part (called Apoenzyme) and non-protein part (called Cofactor), and its activity require these two components. These cofactors may be prosthetic group or coenzyme. a. Prosthetic group is a cofactor tightly bound to enzyme protein as in case of FAD or some metals. So, it is not easy to separated from the enzyme without its destruction. b. Co-enzymes is cofactor loosely (weakly) bound to the enzyme apoprotein. Examples; NAD, FAD and coenzyme-A. 4  11/15/2023 How enzymes accelerate chemical reactions  Enzymes accelerate the rate of reaction by decreasing the energy of activation.  Activation energy: The amount of energy needed to convert a substance (substrate) from ground state to transition state.  The activation energy is very much less for a reaction in presence of enzyme than in non-enzyme catalyst.  Therefore, enzymes are more efficient than non-enzyme catalyst.  Enzyme catalyzed reactions are highly efficient, proceeding from 103 - 108 times faster than uncatalyzed reactions. Enzymes Lower Activation Energy 5  11/15/2023 Nomenclature of Enzymes 1- The International Union of Biochemistry (IUB) classifies the enzymes by giving each enzyme a specific number; this number is called enzyme commission numerical code (EC). It contains 4 digits: EC 1. EC 2. EC 3. EC 4. EC 5. EC 6. First digit = class number. There are 6 classes of enzymes. Second digit = functional group upon which the enzyme acts e.g. -OH, -CHO. Third digit = cofactor e.g. NAD+. Fourth digit = substrate of the enzyme. e.g. Alcohol dahydrogenase: E.C. 1.1.1.1 R-CH2OH + NAD+ R-CHO + NADH + H+ E.C (1): Class of enzyme: oxidoreductase. E.C 1.(1): Group upon which the enzyme acts is: CH2 OH. E.C 1.1.(1): The coenzyme is NAD+. E.C 1.1.1.(1): Alcohol is the substrate. 6  11/15/2023 2- Trivial names (e.g. trypsin, pepsin), which give no indication of the function of the enzyme, or reaction catalyzed by the enzyme, are commonly used. In some cases the trivial names are the names of substrates with the suffix -ase added (e.g., carboxypeptidase, ribonuclease). substrate enzymes products lactose lactase glucose + galactose maltose maltase Glucose cellulose cellulase Glucose lipid lipase Glycerol + fatty acid starch amylase Maltose protein protease Pepton + polypeptide 7  11/15/2023 Factors affecting enzymes activities 1. Substrate Concentration  The rate of an enzyme-catalyzed reaction increases with increasing substrate concentration until maximal velocity (Vmax) is re ached (at substrate saturation).  For most enzymes, the plot of initial reaction velocity (vo) against substrate concentration [S] gives hyperbolic curve (blue curve).  For allosteric enzymes show, the plot of V0 against substrate concentration gives sigmoidal curve (green curve). Factors affecting enzymes activities 2. Enzyme Concentration  At enough substrate concentration, the enzyme velocity increases with increasing enzyme concentration.  This is because as the enzyme concentration increases, the number of active sites increase and the interactions between enzyme molecules and substrate increase leading increasing of enzyme velocity.  At certain point, the substrate molecules decline or finished leading to decreasing or stopping of enzyme velocity. 9  11/15/2023 Factors affecting enzymes activities 3. Temperature  The reaction velocity increases with increasing temperature until reach maximum velocity at optimum temperature.  Further elevation of the temperature over optimum causes a decrease in reaction velocity as a result of temperature- induced denaturation of the enzyme.  The optimum temperature for most human enzymes is 35°C to 40°C. Human enzymes start to denature at temperatures above 40°C.  But thermophilic bacteria found in the hot springs have optimum temperatures of 70°C. Factors affecting enzymes activities 4. pH  At optimum pH of the enzyme, the enzyme active site and substrate are in optimum ionization state and so can interact with each other giving maximum velocity of the enzyme.  Decreasing or increasing pH below or over optimum changes the ionization state of both enzyme and substrate leading to decrease interaction between the and the velocity decrease.  Extremely high or low pH leading to enzyme protein denaturation and so its inactivation.  The optimum pH differ from enzyme to another as in the corresponding curves. 10  11/15/2023 Clinical Enzymology  Plasma enzymes can be classified into two major groups: 1. Small group of enzymes are actively secreted into the blood by certain cell types and have specific physiologic role in the blood. For example, the liver secretes zymogens (inactive precursors) of the enzymes involved in blood coagulation. 2. Large number of enzyme are released from cells during normal cell turnover. These enzymes function intracellularly and have no physiologic use in the plasma.  In healthy individuals, the levels of these enzymes are fairly constant and Increased plasma levels of these enzymes may indicate tissue damage. Clinical Enzymology Alteration of plasma enzyme levels in disease states Many diseases that cause tissue damage result in an increased release of intracellular enzymes into the plasma. The activities of many of these enzymes are routinely determined for diagnostic purposes in diseases of the heart, liver, skeletal muscle, and other tissues. The level of specific enzyme activity in the plasma frequently correlates with the extent of tissue damage. So, determining the degree of elevation of a particular enzyme activity in the plasma is often useful in evaluating the prognosis for the patient. 11  11/15/2023 Clinical Significance of enzyme estimation Assay of serum activity of a selected enzyme can 1) Help in early detection of a disease 2) Help in following up the response to drugs in a disease Enzymes in clinical diagnosis 1) Enzymes can act as diagnostic markers for some diseases. 2) Enzymes can also be used as reagents for various biochemical estimations and detections 23 Enzymes are used as diagnostic markers in different diseases 1) Myocardial Infarction 2) Liver diseases 3) Muscle diseases 4) Bone diseases 5) Cancers 6) GI Tract diseases 12  11/15/2023 Isoenzymes Isoenzymes (isozymes) are different molecular forms of enzymes that may be isolated from the same or different tissues. Isoenzymes characteristics They catalyse the same reaction They have different polypeptide chains. They have different affinities to the substrate. They are affected differently by the different activators and inhibitors. Lactate dehydrogenase (LD) 5 isoenzyme 1- Serum enzymes in Acute 2- Serum enzymes in Liver Myocardial Infarction diseases Enzyme assays routinely carried out for the Enzyme assays routinely carried out for the diagnosis of Acute Myocardial Infarction are: diagnosis of Liver diseases are: (I) Enzymes that Reflect Damage to Hepatocytes A)Creatine Phospho kinase (CPK) A) Alanine transaminase (ALT) B) Aspartate transaminase (AST) B) Aspartate transaminase (AST) (II) Enzymes that reflect Cholestasis C) Lactate dehydrogenase (LDH) HD4 (LDH1) A)Alkaline phosphatase B) 5'-nucleotidase 26 13  11/15/2023 Enzymes as diagnostic reagents Enzyme Used for testing Urease Urea Uricase Uric acid Glucose oxidase Glucose Cholesterol oxidase Cholesterol Lipase Triglycerides Enzymes as therapeutic agents Enzyme Therapeutic Application Streptokinase/Urokinase Acute MI, Pulmonary embolism, DVT(Deep vein thrombosis) Trypsin, lipase and amylase Pancreatic insufficiency Hyaluronidase Enhanced local anesthesia and for easy diffusion of fluids Papain Anti inflammatory Chymotrypsin Pain killer and Anti inflammatory Serratopeptidase Pain killer and Anti inflammatory Alpha- 1 Antitrypsin Emphysema 14  11/15/2023 Enzymes as tumor markers Enzyme Disease Serum acid phosphatase Prostate cancer Serum Alkaline phosphatase Metastasis in liver, jaundice due to carcinoma in the head of pancreas, osteoblastic metastasis in bones Serum LDH Advanced malignancies and Leukemias Β- Glucuronidase Cancer of urinary bladder Leucine Amino Peptidase (LAP) Liver cell carcinoma 15