Mechanisms of Enzymes

Transcription

1 Mechanisms of Enzymes Presented by Dr. Mohammad Saadeh The requirements for the Pharmaceutical Biochemistry I Philadelphia University Faculty of pharmacy

2 How enzymes work * Chemical reactions have an energy barrier separating the reactants and the products. This barrier, called the free energy of activation (Figure 5.4). * To react the molecules of reaction they must contain sufficient energy to overcome the energy barrier of the transition state. * An enzyme allows a reaction to proceed rapidly by providing an alternate reaction pathway with a lower free energy of activation (Figure 5.4). * The enzyme does not change the free energy of the reactant, products and the equilibrium of the reaction.

3 How enzymes work

4 Enzyme enhance the transition state is formed (decrease the activation energy) that can be converted to product and accelerates the reaction.

5 Chemical modes of enzymatic catalysis A. Polar amino residues in active sites Ionizable group found in the active sites. Examples, Histidine is protons accepter or donor. Aspartate, glutamate and lysine have negative or positive charges, can serve as sites by ionic bond.

6 Diffusion-controlled reaction * A reaction that occurs with every collision between reactant molecules is termed a diffusion-controlled reaction. * The apparent second-order rate constant (k cat /k m ) for six very fast enzymes are listed in (table 6.4)

7 Q. From the following data for a hypothetical enzyme, choose the most effective substrate, Explain. The substrate is? Substrate Km Kcat Kcat/Km (M -1 S -1 ) A X X10-3 B 1 2 X X10 2 C 10 2 X D X X10 3 (highest value) The substrate is D because have the highest Kcat/Km value

8 Mechanisms of Enzymes Enzyme mechanisms including kinetic experiments, protein strucural studies, and studies of nonenzymatic model reactions. 1. The terminology of mechanistic chemistry A. Nucleophilic substitutions Nucleophilic= species is electron-rich. Electrophilic= species is electron-poor.

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10 Mechanisms of Enzymes B. Cleavage reactions Covalent bonds can be cleaved in two ways-either both electrons can stay with one atom, or one electron can remain with each atom. C. Oxidation - reductions reactions Loss of electrons = oxidation Gain of electrons = reduction

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12 Structure of chymotrypsin, a serine protease Chymotrypsin (Mr 25,000) is a protease enzyme that catalyzes the hydrolytic cleavage of peptide bonds. Chymotrypsin is specific for cleavage of peptide bonds adjacent to aromatic amino acid residues. Chymotrypsin, example of the use of both covalent catalysis and general acidbase catalysis.

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14 chymotrypsin

15 chymotrypsin

16 chymotrypsin

17 chymotrypsin

18 chymotrypsin

19 chymotrypsin

20 chymotrypsin

21 Hexokinase A hexokinase can be phosphorylates hexoses (glucose) forming hexose phosphate (glucose-6-phosphate). ATP and ADP each bind to the enzymes as a complex with mg +2. Hexokinase, example of induce fit. When glucose is not present, the enzyme is in an inactive conformation with the active-site chains out of position for reaction. When glucose and Mg. ATP bind enzyme conformation change to the catalytically active form.

22 Enolase Enolase (phosphopyruvate hydratase), example of acid-base catalysis and metal ion catalysis.

23 Lysozyme

24 Lysozyme Lysozyme Bacteria cell wall polysaccharide (peptidoglycan). Lysozyme catalyzes hydrolytic cleavage of glycosidic bond between C-1 of (N-Acetylmuramic acid ) MurNAc and oxygen atom at C-4 of a GlcNAc residue involved in glycosidic bond.

25 Lysozyme 1. Lysozyme catalyzes the hydrolysis of polysaccharides, especially those that make up the cell wall of bacteria. 2. Many secretions such as tears, saliva, and nasal mucus contain lysozyme activity to help prevent bacterial infection. 3. Lysozyme catalyzes hydrolytic cleavage of glycosidic bond between C-1 of (N-Acetylmuramic acid ) MurNAc residue and oxygen atom at C- 4 of a (N-Acetylglucosamine) GlcNAc residue (Figure 6.17). 4. Lysozyme accommodates six saccharide residues at six sites (designated A through F). 5. Molecules of polysaccharides binds to lysozyme, MurNAc residues bind to sites B, D, and F. at site D MurNAc distorted into half-chair conformation (figure 6. 19). 6. Two ionic amino acid residues, Glu-35 and Asp-52,are located in C-1 of the distorted sugar molecule in the D binding site. 7. The optimum ph of lysozyme is near 5, between pka of Glu-35 (6.5) and pka of Asp-52 (3.5).

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27 Lysozyme

28 Lysozyme Bacteria cell wall polysaccharide (peptidoglycan). Lysozyme catalyzes hydrolytic cleavage of glycosidic bond between C-1 of (N-Acetylmuramic acid ) MurNAc and oxygen atom involved in glycosidic bond.

29 Lysozyme Bacteria cell wall polysaccharide (peptidoglycan). Lysozyme catalyzes hydrolytic cleavage of glycosidic bond between C-1 of (N-Acetylmuramic acid )MurNAc and oxygen atom involved in glycosidic bond.

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