CC2 LEC

avatar
Created by
As Tï

Cards (381)

  • Factors that influence Enzymatic Reaction
  • Enzymes
    • Proteins that catalyze chemical reactions
    • Biocatalysts that make chemical reactions faster
  • Catalytic Mechanism of Enzymes
    1. A chemical reaction may occur spontaneously if the free energy or available kinetic energy is higher for the reactant than the products (lower energy)
    2. Activation Energy→ reactants have enough energy to break their bond and collide to form new bond (bond between enzyme and substrate)
  • Enzymatic Reaction can be: First-Order Kinetic – the rate of reaction is directly proportional to the substrate concentration. Zero-order kinetic – the rate of reaction is dependent on the enzyme concentration. Saturation kinetic - even if we add more reactant to the reaction once the counterpart reactant has been saturated, it will no longer result in a fast reaction
  • Co-factors are non-protein entities that must bind to an enzyme for a reaction to occur. Activators are made up of inorganic cofactors to alter the spatial configuration of the enzyme, e.g., metallic→ Ca, Fe, Mg, Mn, Zn, K, non-metallic→ Br, Cl. Coenzyme are organic cofactors to serve as a second substrate to enzyme reaction, e.g., Vitamins, AD (nicotinamide adenine dinucleotide); bridge between the enzyme and the substrate
  • When the cell is injured, the enzyme will escape
  • Temperature increase results in an increase in reaction because the molecules' movements are more active in colder temperatures
  • Enzymes are highly specific
    • Absolute Specificity→ the enzyme combined with only one substrate and catalyzed single reaction
    • Group Specificity→ the enzyme combined with all substrates with a particular chemical group
    • Bond Specificity→ the enzyme combined with all substrates with a particular chemical bond such as hydrogen bond, etc.
    • Stereoisometric Specificity→ the enzyme combined with all substrate with the presence of optical isomer (mirror image)
    • Fast chemical reaction is the product of enzyme substrate complex
  • Reactants
    1. Enzymes
    2. Active site - substrate binds the enzyme
    3. Allosteric site – free of water
    4. Substrate
  • Factors that influence Enzymatic Reaction
    1. Substrate concentration/enzyme concentration
    2. Ph- 7-8pH
    3. Temperature
    4. Co-factors
  • Enzymes are essential to physiologic functioning
    • Hydration of CO2 maintains the pH of the blood
    • Nerve conduction in the brain, e.g., CK-BB
    • Muscle contraction for muscular contraction, e.g., CK-MM
    • Nutrient degradation in the GIT, e.g., aminase, lipase
    • Energy use
    • Cardiac enzymes in the heart
    • Liver enzymes in the liver
    • Skeletal muscle enzymes
  • Extreme pH can result in damage or breakdown of enzymes. Acid – ACP, Base – ALP
  • Temperature coefficient – every 10°C increase results in a two-fold increase in the activity of the enzyme. 40-50°C→ start of denaturation of enzyme. 60-65°C→ inactivation of enzyme. 37°C (25 or 30°C) → optimum temperature of enzymatic reaction
  • Storage
    1. Cold temperature can cause reversible inactivation of enzyme
    2. -20°C - long term preservation
    3. 2-8°C - general preservation
    4. Room Temperature - some enzymes are cold labile. Ex. LD (lactate dehydrogenase), LD4 and LD5
    5. Repeated thawing- putting in and out the enzyme forms the freezer can damage it
  • Lactescence/Milky specimen

    Increasing concentration, low measurement. Chylomicrons
  • Isoenzymes
    • Enzymes with the same catalytic function but different physical properties
    • CKBB (CK1) - brain
    • CKMM (CK3) - muscle
    • Differentiated by: Electrophoretic mobility, Solubility, Resistance to inactivation
  • Types of Enzymes
    • Oxido-reductase - catalyze an oxidation-reduction reaction between two substrates
    • Transferase - catalyzed the transfer of a group other than hydrogen from one substrate to another
    • Hydrolase - catalyzed hydrolysis of various bonds
    • Lyase - catalyzed removal of the groups from substrate without hydrolysis; the product contains double bonds
    • Isomerase - catalyzed the interconversion of geometric, optical, or positional isomers
    • Ligases - catalyze the joining of the two substrate molecules, coupled with breaking of the pyrophosphate bond in adenosine triphosphate (ATP) or a similar compound
  • Coenzyme

    • Organic cofactors; to serve as a second substrate to enzyme reaction
    • Ex. Vitamins, AD (nicotinamide adenine dinucleotide); bridge between the enzyme and the substrate
  • Enzymes
    Proteins (Amino Acid)
  • Inhibitors interfere with the chemical reaction
    1. Types of Inhibition: Competitive Inhibition - the target of the inhibitor is the active site
    2. Non-Competitive Inhibition - the target is the allosteric site
    3. Uncompetitive Inhibition - the target is the E.S complex
  • Enzymology Part ii
    1. Creatine Kinase - Associated with ATP generation in the contractile system (muscle tissue)
    2. Function: in the muscle cells, it stores Creatine phosphate that is important in ATP production
    3. Tissue sources: Skeletal muscle, Heart muscle, Brain tissue
    4. Isoenzymes: Dimer with 2 subunits
    5. CK-1 - <1% aka CK-BB - brain type
    6. CK-2 - <6% CK-MB - hybrid type
    7. CK-3 - 94-100% CK-MM - muscle type (blood)
    8. Isoenzymes: by electrophoresis, we can differentiate CK
    9. CK-1= <1% - most anodal (anode)
    10. CK-2= <6%
    11. CK-3= 94-100% - most cathodal (cathode)
  • Enzymatic Reactions
    1. 2 Methods in measuring the enzymatic reactions: Fixed-time - single measurement of the sample
    2. Continuous-Monitoring/Kinetic Assay - multiple measurement of enzyme activity
  • Enzyme structure
    • Primary Structure - specific sequence of amino acid
    • Secondary Structure - polypeptide chain twisting
    • Tertiary Structure - folding of the polypeptide chain/secondary structure
    • Quaternary Structure - combination of tertiary structure
  • Nomenclature-naming developed by the E.C. (Enzyme Commission)
  • Alter the spatial configuration of enzyme
    1. Metallic: Ca, Fe, Mg, Mn, Zn, K
    2. Non-metallic: Br, Cl
  • Hemolysis

    Destruction of RBC; K(Potassium) L(LDH) A(AST, ACP, aldolase) M(magnesium) P(phosphate) enzyme will increase during hemolysis
  • Rise in CK-MB
    1. 8 hrs
  • CK-1

    Less than 1%, most anodal (anode)
  • CK-BB

    • Seldomly found in plasma, short half-life, high molecular size
  • CK-3

    94-100%, most cathodal (cathode)
  • Normalization of CK-MB
    48-72 hrs
  • Macro CK

    Found midway between MM and MB
  • Oliver-Rosalki Assay

    Measurement of Creatine Phosphate + ADP to Creatinine + ATP
  • Methods used for the measurement of Isoenzymes of CK
    1. Electrophoresis
    2. Ion Exchange Chromatography
    3. Antibodies
    4. Immunoassay
  • Lactate dehydrogenase catalyzes the interconversion of lactic acid and pyruvic acid
  • For storage, CK can last up to 7 days at 4 degrees Celsius and 1 month at -20 degrees Celsius
  • CK-MM
    • Major fraction in serum; skeletal muscle
    • Elevated in hypothyroidism, muscle activity, IM injection
  • Peak in CK-MB
    12-24 hrs
  • CK-2

    Less than 6%
  • CK-MB
    • 20% of cardiac tissue contains CK-MB, specific to heart muscle, indicator of myocardial damage