Biochemistry I Spring/Summer 2024 Lecture 5 - 2024 PDF

Biochemistry I Spring/Summer 2024 Lecture 5 April 9, 2024 Textbooks: Mark’s Basic Medical Biochemistry 5th Edition And Harper’s Illustrated Biochemistry 30th Edition Read Chapters 8-11 in Mark’s Basic Medical Biochemistry Please read chapters 8-11 Tentative list o...

Biochemistry I Spring/Summer 2024 Lecture 5 April 9, 2024 Textbooks: Mark’s Basic Medical Biochemistry 5th Edition And Harper’s Illustrated Biochemistry 30th Edition Read Chapters 8-11 in Mark’s Basic Medical Biochemistry Please read chapters 8-11 Tentative list of topics and chapters for the rest of the semester: Mark’s Basic Medical Biochemistry, Reading: Chapter 8 enzymes as catalysts Chapter 9 regulation of enzymes Chapter 10 relationship between cell biology and biochemistry Chapter 11 cell signaling 12-18 gene expression and protein synthesis Chapters 19-28 Carbohydrate metabolism. Fuel oxidation and generation of ATP Enzymes are catalysts that vastly increase the rate (velocity) of a chemical reaction Rate(cat.)/Rate (uncat.) = 106 - 1014 Speed (velocity), specificity, and regulation Glucokinase (liver, pancreas) – only acts on glucose (specificity), regulation- synthesis of glucokinase is induced by insulin. Glucokinase Classification : a transferase, in particular, a kinase- transfer of phosphate group Binding of a substrate E + S ES Conversion of substrate to product ES → EP Release of product EP E+P Function, specificity and speed depend on the sequence of amino acids that comprise the tertiary structure Substrate recognition site or coenzymes For example ATP in glucokinase  Glucose-binding site in glucokinase. A. Glucose, shown in red, is held in its binding site by multiple hydrogen bonds between each hydroxyl group and polar amino acids from different regions of the enzyme amino acid sequence in the actin fold (see Chapter 7). The position of the amino acid residue in the linear sequence is given by its number. The multiple interactions enable glucose to induce large conformational changes in the enzyme (induced fit). (Modified from Pilkis SJ, Weber IT, Harrison RW, et al. Glucokinase: structural analysis of a protein involved in susceptibility to Aspartic acid pKa 3.86 diabetes. J Biol Glycine Chem. 1994;269(35):21925– Glutamic acid pKa 4.25 21928. https://creativecommons.org/lic enses/by/4.0/) B. Enzyme specificity is Asparginine illustrated by the comparison of Aspartic acid galactose and glucose. Galactose differs from glucose only in the position of the –OH group shown in red. However, it is not phosphorylated at a significant rate Side note: glucose and galactose are both by the enzyme. Cells therefore require a separate galactokinase for the D-hexoses but differ at carbon 4 : epimers metabolism of galactose. Glucokinase (liver, pancreas) and hexokinase (other tissue) E+S ES ES* EP E+P Enzymes increase the rate of reaction by decreasing activation energy Enzyme catalytic startegies: electronic stabilization of transition state complex, acid-base catalysis, This again, is accomplished by the sequence of amino acids Energy diagram showing the energy levels of the substrates as they progress toward products in the absence of enzyme. The substrates must pass through the high-energy transition state during the reaction. Although a favorable loss of energy occurs during the reaction, the rate of the reaction is slowed by the energy barrier to forming the transition state. The energy barrier is referred to as the activation energy.  A. General acid-base catalysis: an amino acid side chain either donates a proton (acid) or accepts a proton (base) example: chymotripsin is a serine protease. Histidine 57 acts as a generl base and abstracts a proton from serine 195. Protonated histidine 57 donates the proton (general acid catalysis) to the product. chymotripsin B. Covalent catalysis. Ex chymotripsin: deprotonated serine attacks the carbonyl group of a peptide bond forming a temporary covalent bond or tetrahedral intermediate https://www.youtube.com/watch?v=Z0wqKJfd xgE Lecturio Medical video on serine proteases C. Metal Ion Catalysis D. Catalysis by approximation Or catalysis by proximity and orientation PDC Coenzymes: non-protein organic cofactors covalently or noncovalently bound to an enzyme or eventually become incorporated into the product. Tightly bound coenzymes are called prosthetic groups. Activation-transfer coenzymes: catalysis by covalent bond formation with a substrate to activate it for additional reactions PDC has 5 coenzymes Coenzyme A Pyruvate carboxylase: 1st step of gluconeogenesis bicarbonate ion is phosphorylated by ATP and thus is activated for decarboxylation, which generates free CO2 Enzyme Kinetics ⎯⎯ k1 E + S ⎯ → ⎯ ⎯⎯ ES ⎯⎯ k2 → EP ⎯⎯ k3 ⎯⎯ →E + P k−1 k−2 k−3 Simplified ⎯⎯ k1 E + S ⎯ → ES ⎯⎯→ ⎯ k cat E+P : k−1 V0 = [product]/time or –[S]/time Mol/Ls Keep enzyme concentration constant- vary substrate concentration Low [substrate] High [substrate]  ⎯⎯k1 E + S ⎯⎯ → ES ⎯⎯→ k cat E+P k−1  The Michaelis-Menton Equation kcat [E]t [S] or Vmax S At saturation: v= v= K M + [S] K M + S kcat [E]t = Vmax Velocity (mol/Ls) Interpreting KM, kcat, and kcat/KM (2 of 2) Diffusion-limit for kcat/KM values is 108 to 109 (mol/L)−1 s−1. Enzymes reaching this limit are said to be “perfect”  The ratio kcat/KM is a convenient measure of enzyme efficiency and substrate specificity For example: