enzymes l.
Download
Skip this Video
Loading SlideShow in 5 Seconds..
Enzymes PowerPoint Presentation
Download Presentation
Enzymes

Loading in 2 Seconds...

play fullscreen
1 / 20
Leo

Enzymes - PowerPoint PPT Presentation

169 Views
Download Presentation
Enzymes
An Image/Link below is provided (as is) to download presentation

Download Policy: Content on the Website is provided to you AS IS for your information and personal use and may not be sold / licensed / shared on other websites without getting consent from its author. While downloading, if for some reason you are not able to download a presentation, the publisher may have deleted the file from their server.

- - - - - - - - - - - - - - - - - - - - - - - - - - - E N D - - - - - - - - - - - - - - - - - - - - - - - - - - -
Presentation Transcript

  1. Enzymes • Enzymes are catalytically active proteins that regulate virtually all biological processes. Industrial applications can be found in: • Fine chemical preparations • Food-industry (dairy, starch conversion) • Analytical chemistry and medicine • While the activity of enzyme-mediated reactions is exceptional, it is selectivity of these processes that is both unique and valuable: • Reaction specificity • Regioselectivity: yields one of several structural isomers • Stereoselectivity: consumes/yields one stereoisomer (enantiomer) • Topics for Discussion • Enzyme production and purification schemes • Industrial applications • Enzyme structure and the nature of the catalytic site • Catalytic chemistry and reaction kinetics • Enzyme immobilization and mass transfer

  2. Enzyme Production/Isolation Methods • The structural complexity of enzymes makes their synthetic preparation a formidable task. They are natural products that are isolated/produced from three principle sources. • Isolation from animal organs (hog insulin) • Isolation from plant material (papain) • Microorganism production • Isolation and purification are complicated by the presence of similar proteins and the inherent sensitivity of enzymes to pH, temperature and degradation by other enzymes.

  3. Classification of Enzymes • I. Oxidoreductases: encompass all enzymes that catalyze redox reactions. Name is dehydrogenase whenever possible, but reductase can also be used. Oxidase is used only where 0 is the acceptor for reduction. • 2. Transferases: catalyze the transfer of a specific group such as methyl, amino or phosphate from one substance to another. Name is acceptor group- transferase or donor group-transferase. • 3. Hydrolases: catalyze the hydrolytic cleavage of C-O, C-N, C-C, and some other bonds. Name often consists of the substrate name with the suffix -ase. • 4. Lyases: catalyze the cleavage of C-C, C-O, C-N, and other bonds by elimination. Name is, for example, decarboxylase, dehydratase (elimination of CO and water, respectively). • 5. Isomerases: catalyze geometric or structural rearrangements within a molecule. Different types lead to the names racemase, epimerase, isomerase, tautomerase, or cycloisomerase. • 6. Ligases: catalyze the joining of two molecules, coupled with the hydrolysis of a pyrophosphate bond in ATP or another nucleoside triphosphate.

  4. Applications of Proteolytic Enzymes • Dairy: • Calf rennet (chymosin) is used in the coagulation of milk protein for cheese production, without loss of sensitive components. • Lactase hydrolyzes the principal carbohydrate of milk, lactose. This processes a cheese byproduct and relieves lactose intolerance. • Detergents: • Protein stain removal is facilitated by the hydrolysis of proteins into oligopeptides. Enzyme stability with respect to storage, pH, temperature and bleach are key concerns. • Leather Production: • Proteases are widely used for the soaking and dewooling stages of hide processing in which selective protein degradation results in a softer produce without substantial loss of strength. • Food and Feed: • Starch conversion to high-fructose corn syrup is an important process to the beverage industry.

  5. Starch Conversion • The processing of starch to yield sweeteners is a significant industrial operation. Acid-catalyzed hydrolysis has been largely supplanted by enzyme-mediated hydrolysis due to superiour activity and reduced by-product formation. • Starch contains two polysaccharides, amylose and amylopectin. • Amylose (n approx. 400) • amylopectin • (branched) • Hydrolysis of amylose yields the constituent monomer D-glucose, while the degradation of amylopectin is complicated by its branched structure.

  6. Starch Conversion: High Fructose Corn Syrup • a-amylase degrades amylose to D-glucose, but a second enzyme, glucoamylase is needed to breakdown oligosaccharides derived from amylopectin. • This product can be used as substrate for yeast fermentation to produce ethanol as an alternate fuel source. Much of the glucose produced by starch degradation is isomerized to fructose for use as a low-cost (relative to sucrose) natural sweetener in soft drinks

  7. Starch Conversion: Isomerization

  8. Enzyme “Building-Blocks”: a-Amino Acids • An enzyme is a singular macromolecule with precise monomer sequencing. As with all proteins, the monomers that constitute enzymes are a-amino acids: • The zwitterionic character results from the potential for proton donation from the carboxylic acid group to the basic amino functionality of the molecule. • Condensation of amino acids yields biological oligomers/polymers known as peptides, which if catalytically active are enzymes. • Degrees of polymerization • from 60 to 1000 are • known.

  9. Enzyme “Building-Blocks”: a-Amino Acids • There are twenty amino acids that occur commonly as constituents of most proteins. With but one exception (proline) all a-amino acids have the same general structure, differing only in the substituent R. • With the exception of glycine (R=H), the • a-amino acids have at least one • asymmetric carbon atom that exists in • the S-configuration. • The backbone structure of peptides derived from the a-amino acids is capable of hydrogen-bonding to yield highly ordered chain conformations. • The substituents, R, range from non-polar aliphatics and aromatics to polar alcohols, amines and carboxylic acids. • The nature of a substituent affects enzyme conformation as well as the chemistry by which a reaction is catalyzed.

  10. Common a-Amino Acids

  11. Common a-Amino Acids

  12. Structure of Enzymes • Enzymes have genetically mandated and unique amino acid sequences: • Although only a small subset of the amino acids within an enzyme may engage the reactant(s), all enzyme constituents are needed for catalytic activity. Enormous molecule size generates: • sufficient local-controlled flexibility • precise three dimensional arrangements • In spite of the tremendous structural complexity of enzymes, reactions derived from their reactive functional groups are similar to the acid-base and metal-mediated processes you have already studied.

  13. Primary Structure of Enzymes • The complex structure of enzymes can • be discussed at different levels, the • simplest of which is the covalent • structure, or primary structure. • The most important aspect is the • amino acid sequence, shown here • for lysozyme. • However, peptide bonds • alone do not define primary • structure, as disulfide bonds • between cysteine residues • crosslink different parts of the peptide chain.

  14. Secondary Structure of Enzymes • Macromolecules which lack polar functional groups assume random coil configurations in solution that are dictated by polymer-solvent interactions. • Peptides on the other hand have • restricted rotation about the • carbonyl-nitrogen bond of the amide • linkage, thereby “locking” one site of • potential backbone flexibility into • the Z-conformation. • The description of enzyme structure in terms of ordered domains is referred to as the secondary structure. Three peptide conformations are most commonly assumed: • random coil conformation • a-helix • b-pleated sheet

  15. Secondary Structure of Enzymes • A common peptide backbone configuration is a right-handed a-helix. • A helical conformation of the enzyme is generated through hydrogen-bond interactions between the amide N-H of • one residue and the carbonyl oxygen four residues away. • The side-chain groups are positioned on the outside of • the helix.

  16. Secondary Structure of Enzymes • A b-structure, or pleated sheet is generated by hydrogen-bonding interactions between peptide chains (or a different part of the same chain). • A peptide chain adopts an open, zigzag conformation

  17. Tertiary Structure of Enzymes • The complete three dimensional structure of a protein is called its tertiary structure. It is an aggregate of a-helix, b-sheet and random coil and other structural elements that is governed by non-covalent interactions. • While covalent (peptide and disulfide • bonds) interactions define the primary • structure, non-covalent interactions • determine the tertiary structure • of proteins: • Hydrogen bonding • Van der Waals interactions • Electrostatic interactions • Note that amino acid sequencing, as • defined by the primary structure, • determines the extent of these non-covalent interactions, and defines the secondary and tertiary structure of the enzyme.

  18. Tertiary Structure of Enzymes • In solution an enzyme adopts a lowest-energy conformation which, owing to the uniqueness of the primary structure, is very precise. • The polarity of the amino acid functional groups dictates the affinity of particular peptide sequences for water. Hydrophilic sequences favour positions on the surface of the enzyme, while hydrophobic sequences are found in the internal regions of the protein. • This behaviour is analgous to micelle formation in surfactants.

  19. Quaternary Structure of Enzymes • Many enzymes consist of more than one polypeptide chain (or subunit) that aggregate to confer catalytic activity. • In some enzymes the subunits are identical, in other cases they differ in sequence and structure. • The description of subunit arrangement in such enzymes is called the quaternary structure. • A typical enzyme is not an entity completely folded as a whole, but may consist of apparently autonomous or semi-autonomous folding units called domains. • Functional domains (those providing catalytic activity) can be regions that fold independently • The active site of lysozyme (slide 11) is believed to be the cleft between two distinct domains.

  20. Summary of Enzyme Structure • Enzymes are catalytically active macromolecules comprised of a specific sequence of a-amino acids. • The sequence of amino acids through peptide bonds and chain crosslinking through disulfide bonds of cysteine residues is the primary structure of the enzyme. • Peptide sequences can form ordered subunits through hydrogen bonding interactions. These include the a-helix, b-sheet. Random coil conformations predominate in the remaining peptide sequences. These comprise the secondary structure of the enzyme. • Non-covalent interactions between the elements of the secondary structure generate the very specific overall conformation of the enzyme, called the tertiary structure. • Where more than one peptide chain aggregates to generate the active enzyme, a quaternary structure is defined. • The structure of an enzyme represents the lowest-energy conformation of the macromolecule, which will spontaneously form given the appropriate primary structure.