BIOL 541 1st Edition Lecture 11 Outline of Last Lecture I. Co enzymes and co factors. Outline of Current Lecture II.Molecular mechanisms of catalysis.III. Six major groups of enzymes.Current LectureBiochem Biol 541: Lecture 11:Glomerulization of enzymes- metabolon.3 examples of molecular mechanisms of catalysis:Lysozyme: This is found in tears and egg whites. It is anti-bacterial in nature and digests bacterial cell wall. It has specificity for polysaccharide that alternates between acetyl glucosamine and OH that replaces thehetero polysaccharide: 2 sugars forms NAG and NAM. It is a relatively small protein of 129 amino acids and a molecular weight of 14,600 Da. It is single chain globular in structure and stabilized by intra chain disulphide bonds. It has an overall cleft which is the substrate binding site- places for sugars. Amino acidsfrom lysozyme can make contact with the polysaccharide chains.There are non covalent bonds that decrease the free energy: all except those for binding of 4th sugar. Chain fits into cleft. D- sugar is in the ½chair conformation i.e. stressful and high energy conformation. It is not favorable except it decreases free energy from 5th residue in order to drive the reaction: therefore, termed as “energy leaver”. There is also reduced fit (allosterism) Try 62 moves in 1 A upon substrate binding. Cleavage occurs at ½ chair sugar and cleavage proceeds via lactone derivative that forms ½ chair. ½ chair is intermediate structure (active sites and catalysis). Inactive site, protonated GLU acid and ionized ASP acid occurs at pk3. However, local environment increases pk3 value so that both can be protonated. But in tears ( since These notes represent a detailed interpretation of the professor’s lecture. GradeBuddy is best used as a supplement to your own notes, not as a substitute.extracellular fluid is acidic) has pH 5.0. GLU acid has a value more than that and ASP has a value less thanthat. Therefore, GLU acid is protonated and ASP acid is ionized. H from the GLU acid acts as acid and donates H.Therefore, this is an example of acid catalysis and destabilization. Glycidic bond is attacked as H is attacked. N resonates as ½ chair which is facilitated by ½ chair conformation: ASP acid stabilizes (+) charge. Water comes in, GLU acid grabs H back, thereby the C and ½of the polysaccharide leaves. Lysozyme binds and cleaves it. Hydrophilic residue and distortion of substrate and acid catalysis are happening.Carboxy peptidase A: This nibbles at carboxy terminus except for basic amino acid (carboxy peptidase B attacks this and Carboxy peptidase C acts on proline). Zn proteases acts as co- factor. Pancreatic enzyme like lysozyme is globular 307 amino acid with one Zn ion. Active site has non polar pocket and ARG for selectivity and attracts the caboxy terminus which is (-). Zn is co-ordinated by 2 HIS and GLU acid. HIS kelates Zn and carboxy peptidase A. GLU acid and TYR peptide bond cleavage occurs. Dramatic induced fit of 2 A moving in: ZN positioned such that it pulls on oxygen. Carbonyl of the alternate amino acid is polar and Zn pulls and the pole extends to cleave peptide bond and release terminal amino acid. Mechanism of cleavage is either by attack of GLU acid 270 or by water. X- is the terminal of the amino acid.Zn pulls c=o, and then GLU 270 and TYR 248 start. GLU acid attacks c=o C and Zn increases polarity. C becomes more (+) and COOH which is (-) attracks it rupturing = bond because C can not be more than 4 bonds. Peptide formed as c=o comes back as terminal amino acid is released. H binds to amino group for acid hydrolysis. Therefore, both covalent catalysis and general catalysis occurs. The enzyme is released and regenerated. H reprotonates TYR and OH brakes ester bond of GLU acid.Acid base catalysis by activated water: c=o is pulled by Zn, TYR, and GLU acid. GLU acid pulls on H of water to form OH. OH is going to attach c=o and break the peptide bond. Base catalysis occurs, H from TYR breaks OH, and H binds to terminal amino group. Therefore term used is acid catalysis. H dissociates GLU acid and reprotonate TYR and active site is regenerated.Chymotrypsin: This is a pancreatic enzyme which begins as a single peptide called chymotrypsinogen (precursor common for proteases). During storage there should be no need for activation. Zymogen is the term for the precursor of any enzyme. When protein rich meal is ingested pancreas secretes enzymeand is activated in duodenum. 245 amino acids are clipped between 13 and 16: 146 and 149. Since the bonds are held by disulphide bonds, clippage exposes ISO LEU 16. Amino group in peptide bond with amino acid 15 forms amides and not ionized. Free amino acid is therefore protonated. (+) charge is attracted to ASP 194 and interaction results in change in conformation and pulls a MET (hydrophobic amino acid) and forms a pocket. Chymotrypsin cleaves amino acid which is hydrophobic as specificity exists due to MET.Active site: Non polar pocket cleaves peptide. TYR is bound and cleaves on amino side. Oxygen of SER 195 attacks the bond. SER is not ionized as pk3 value is 13. Local environment: ASP (-) charge pulls H off and creates (-) charge on oxygen which becomes reactive and swings down to attack the peptide bond. ASP 102, HIS 57, and SER 195 constitute charged relay system.Covalent catalysis: Ionized oxygen can attack c=o. Electrons go into ring and electrons bind H from SER. (-) SER attracts c=o. (-) Oxygen adds to (+) C and breaks double bonds of C.The peptide is covalently bound to enzyme. Rearrangement occurs where electrons flow back and free H. Oxygen needs are satisfied. H attacks peptide bond and joins amino group. Therefore, the process is acid catalysis. C=O is reformed. The cleaved peptide bond: ½ peptide is left and ½ is covalently bound to enzyme. ASP acid pulls on electrons. Activated water releases OH group that attacks the ester bond. ½ peptide leaves, electrons flow back and SER gets H back. Electrons go back and forth as it binds to hydrophobic acid. Therefore, covalent catalysis, acid catalysis and basic catalysis occurs. Trypsin is homologous to this: ASP acid in the binding site instead of MET. Here (+) charged amino acid is cleaved forming specificity changes to form binding pockets.Thiol proteases: Papain (meat tenderizer). It has the same basic charge relay system where CYS has been substituted for SER. S and