Ch395G Fall 2009: Lecture 16Enzymatic CatalysisAnd ControlAnd ControlEnzyme Catalysis• Lysozyme as an example – look for how rate is enhanced and the methods used to determine mechanistic information.•Biochemical Data•Biochemical Data• Structural Data• Phillips Mechanism• Recent DevelopmentsSummary –What “chemical” mechanisms were employed in Lysozyme?What methods provided the data to work out the current mechanism?CorrectionI had noted earlier that procaryotes do not carry out protein glycosylation and that this resulted in problems for making some effective eucaryoticrecombinant proteins in bacterial hosts.Recently it was discovered the Campylobacter jejuni can do both N- and O- linked glycosylation. The systems for doing this glycosylation have now been transferred to E.coli.Control of Enzymatic ActivityRates of synthesis and degradation (Nobel Prize for the ubiquitination pathway)See Chapter 13 pages 465 - 470for the ubiquitination pathway)Proenzyme (zymogen) conversionPresence of activators and inhibitorsCovalent ModificationAllosteric ControlFrom the National Museum of MedicineUbiquitin tagging of proteins for degradationIrwin Rose (UCSD) one of the 3 Nobel Prize winnersEnzyme Control by Covalent ModificationEnzymes can either by activated or deactivated by phosphorylation.Phosphorylation sites include tyrosine, serine and threonineAllosteric Control of Enzymatic ActivitySee Chapter 10 pp 345-353 for introduction to this topicTwo Different Theoretical ModelsTwo Different Theoretical ModelsSymmetry Model (Monod, Wyman, Changeux 1965)Sequential Model(Koshland 1966)This material was not covered in depth in the lectures and questions will only be asked in exams to the level it was covered in class.Figure 10-31 Heterotropic interactions in the symmetry model of allosterism.Page 349Figure 10-32 The effects of allosteric activator (γ = [A]/kA) and inhibitor (β = [I]/kI) on the shape of the fractional saturation curve for substrate (α = [S]/kR).Page 349Figure 10-34 The sequential model of allosterism.Page 350Sequential ModelFigure 10-35 Sequential binding of ligand in the sequential model of allosterism.Page 351Figure 10-36 The sequential and the symmetry models of allosterism can provide equally good fits to the measured O2-dissociation curve of Hb.Page 351P 465 topFigure 13-6 Schematic representation of the pyrimidine biosynthesis pathway.Page 466Figure 13-5 The rate of the reaction catalyzed by ATCase as a function of aspartate concentration.Page 465Figure 13-7b X-Ray structure of ATCase. T-state ATCase along the protein’s molecular twofold axis of symmetry.Page 467Figure 13-7b X-Ray structure of ATCase. R-state ATCase along the protein’s molecular twofold axis of symmetry.Page 467Figure 13-8 Comparison of the polypeptide backbones of the ATCase catalytic subunit in the T state (orange) and the R state (blue).Page 468Figure 13-9 Schematic diagram indicating the tertiary and quaternary conformational changes in two vertically interacting catalytic ATCase subunits. Page
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