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Enzymatic Mechanisms

We can apply what we learned about catalysis by small molecules to enzyme-catalyzed reactions. To understand the mechanism of an enzyme-catalyzed reaction, we try to alter as many variables, one at a time, and ascertain the effects of the changes on the activity of the enzyme. Kinetic methods can be used to obtain data from which inferences about the mechanism can be made. Obviously, crystal structures of the enzyme in the presence and absence of a competitive inhibitor give abundant information about possible mechanisms.

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Many enzymes have active site serines which act as nucleophilic catalysts in nucleophilic substitution reactions (usually hydrolysis).; One such enzyme is acetylcholine esterase which cleaves the neurotransmitter acetylcholine in the synapse of the neuromuscular junction.; The transmitter leads to muscle contraction when it binds its receptor on the muscle cell surface.; The transmitter must not reside too long in the synapse, otherwise muscle contraction will continue in an uncontrolled fashion.; To prevent this, a hydrolytic enzyme, acetylcholine esterase, a serine esterase found in the synapse, cleaves the transmitter, at rates close to diffusion controlled.; Diisopropylphosphofluoridate (DIPF) also inhibits this enzyme which effectively makes it a potent chemical warfare agent.; An even more fluoride-based; inhibitor of this enzyme, sarin, is the most potent lethal chemical agent of this class known.; Only 1 mg is necessary to kill a human being.

Figure:; sarin

12sarin.gif

 

Other Types of Enzymes

The three enzymes studied above are all hydrolases - enzymes that catalyze the hydrolysis of bonds (either amide or acetal).; This is only one class of six different reaction types that have been categorized by the Enzyme Commission of the International Union of Biochemistry and Molecular Biology.; The six types include:

  • EC1:; Oxidioreductases - oxdiation/reduction reactions (we will discuss these in Chapter 8B)
  • EC2:; Transferases - acyl, glycosyl, 1C, N, O, aldehydes, ketones, etc;
  • EC3:; Hydrolases
  • EC4:; Lyases - cleavage of C-C, C-O, C-N, C-S, etc. bonds
  • EC5:; Isomerases - racemases, epimerases, cis-trans isomerases
  • EC6:; Ligases - form C-C, C-O, C-N, etc bonds
  • Enzyme Nomenclature Database:; Interactive site to search information on enzymes using EC system of nomenclature.
  • BRENDA:; (Brauschweig Enzyme Database) Comprehensive Enzyme Information System
  • KEGG PATHWAY: ; collection of manually drawn pathway maps representing our knowledge on the molecular interaction and reaction networks using KEGG, Kyoto Encyclopedia of Genes and Genomes
  • FMM (From Metabolite to Metabolite) - reconstructs metabolic pathways from one metabolite to another

 

References

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  2. Wang, Y. et al. Crystal structure of a rhomboid family intramembrane protease.; Nature.; 444, 179 (2006)
  3. Freeman, M. Proteolysis within the membrane: rhomboids revealed.; Nature Reviews: Molecular Cell Biology. 5, p 188 (2004)
  4. Borman, S. Much ado about enzyme mechanisms.; C&EN.; pg 35 (Feb 23, 2004)
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  8. Weihofen et al. Identification of Signal Peptide Peptidase, a Presenilin-Type Aspartic Protease. Science,; 296, pp. 2156, 2215,
  9. Vocadlo. D. et al.; Catalysis by hen egg-white lysozyme proceeds via a covalent intermediate.; Nature. 412. pg 835 (2001)
  10. Walsh, C. Enabling the Chemistry of Life.; Great review article on enzymes mechanisms.;; Nature. 409, pg 226 (2001)
  11. Koeller and Wong.; Enzymes for Chemical Synthesis.; Nature 409. pg 232 (2001)
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  13. Huntington et al. Structure of a serpin-protease complex shows inhibition by deformation.; Nature. 407, pg 923 (2000)
  14. New Way to Study Closely related proteins (remodeling proteins to make them more susceptible to inhibition); Science 289. pg 2029 (2000)
  15. Vocadlo et al. Catalysis by hen egg-white lysozyme proceeds via a covalent intermediate.; Nature. 412. pg 835 (2001)
  16. Istan and Deisenhofer. Structural Mechanism for Statin Inhibition of HMG-CoA Reductase. Science. 292, pg 1160 (2001)
  17. Heine et al. Observations of Covalent Intermediates in an Enzyme Mechanism at Atomic Resolution.; Science 294. pg 369 (2001)
  18. Carpenter et al. Structure of dehydorquinate synthase reveals an active site capable of multi-step catalysis.; Nature. 394, pg 299 (1998)
  19. Kohen et al. Tunnel Vision (on why activity of therophilic enzymes (>60oC); is low or absent at mesophilic temperatures (< 40oC) - from reduction of flexibility of thermophilic enzymes at mesophilic temperatures; - quantum tunneling explanation).; Nature. 399, pg 417, 496 (1999)
  20. Finnin et al. Structure of a histone deacetylase homologue bound to the TSA and SAHA inhibitors (and mechanism).; Nature. pg 189, September 1999.
  21. Ondrechen. THEMATICS: A simple computational predictor of enzyme function from structure.; Proc. Natl. Acad. Sci. USA, 98, pg 12473 (2001)