Ch 10.0-.1 Text

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Ch 10.0-.1 Text
2014-10-22 13:30:23
Test Three
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  1. ·         Regulation of enzymes allows them to __. There are five enzymatic forms of control.
    WHat are the five?
    function at proper time and place and allows coordination of activity

    • allosteric
    • multiple forms of enzyme
    • reversible covalent modification
    • proteolytic activation
    • controlling amount of enzyme present
  2. Allosteric
    • o   contain distinct regulatory sites and multiple functional sites. Small signal molecules bind, allowing cooperativity to transduce information.
    • §  Ex: ATCase
    • ·         Catalysis by aspartate transcarbamoylase of the first step is inhibited by CTP through feedback inhibition
  3. Multiple forms of the enzyme
    o   isozymes allow varying regulation of the same reaction at distinct locations or times to meet certain physiological needs in that particular tissue. They are homologous enzymes within a single organism that catalyze the same reaction but differ slightly in structure and KM and Vmax values
  4. Reversible Covalent Modification
    o   involve covalent attachment of a modifying group, usually phosphoryl groups from ATP, which is catalyzed by protein kinases and hydrolyzed by protein phosphatases
  5. proteolytic activation
    • o   the enzyme can be active or inactive; a different strategy is used to irreversibly convert an inactive enzyme into an active one
    • §  Many enzymes are activated by hydrolysis of a few peptide bonds or one in zymogens (proenzymes)
    • ·         Ex: digestive enzymes such as chymotrypsin
  6. Controlling the amount of enzyme present
    this takes place at the level of transcription
  7. ·         _ catalyzes the first step in the biosynthesis of pyrimidines: the __ to form __. This is the __ in the pathway that will ultimately yield pyrimidine nucleotides like CTP
    • Aspartate transcarbamoylase
    • condensation of aspartate and carbamoyl phosphate
    • N-carbamoylaspartate and orthophosphate
    • committed step reaction
  8. ·         Regulation occurs because __.
    o   The rate of the reaction catalyzed by __is fast at __but slows as __. Thus, the pathway continues to do what until what?

    o   This is __, inhibition of an enzyme by the end product of the pathway; it ensures by __ and intermediates are not formed when pyrimidines are abundant
    • CTP inhibits ATCase
    • ATCase 
    • low concentrations of CTP 
    • CTP concentration increases
    • make new pyrimidines
    • enough CTP is present
    • feedback inhibition
    • N-carbamoylaspartate
  9. ·         CTP is structurally different from the substrates of the reaction and must bind to site different from active site, called an __. It is an __
    ·         ATCase consists of __ and can be separated by treatment with a __compound, which reacts with __.
    o   __after reveals the __, which can be separated by __ due to different charge or by __due to different size
    • allosteric or regulatory site
    • allosteric inhibitor
    • separable catalytic and regulatory subunits
    • mercurial 
    • sulfhydryl groups
    • Ultracentrifugation 
    • two subunits
    • ion-exchange chromatography
    • centrifugation
  10. ·         The larger subunit is the __, which displays __activity but is __ and does not display __
    o   Consists of __; there are __
    ·         The isolated smaller subunit can bind __, but has no __ activity, which is why it __
    Consists of __; there are __
    • catalytic subunit
    • catalytic 
    • unresponsive to CTP
    • sigmoidal kinetics
    • three chains
    • two catalytic trimers
    • CTP
    • catalytic
    • regulates
    • two chains
    • three regulatory dimers
  11. ·         ATCase is composed of discrete __ and __, and the interaction of the subunits in the native enzyme produces its __ and __
    Allosteric interactions in ATCase are mediated by __
    • catalytic and regulatory subunits
    • regulatory and catalytic properties
    • large changes in quaternary structure
  12. o   Two catalytic trimers are arranged how? There are significant contacts between the __ and __ subunits: each __ within a __ interacts with a __ within a __
    The __ makes contact with a structural domain in the __ that is stabilized by a __ion bound to __ residues
    • stacked one on top of the other, linked by three dimers of the regulatory chains
    • catalytic and the regulatory subunits
    • r chain
    • regulatory dimer
    • c chain
    • catalytic trimer
    • c chain
    • r chain
    • zinc 
    • four cysteine
  13. ·         To locate the active sites, the enzyme was crystallized in the presence of __, a bisubstrate analog that resembles an intermediate along the pathway of catalysis
    o   __is a potent __ of ATCase. It binds at sites where, each of which contributes __ to the complete enzyme

    • PALA
    • PALA 
    • competitive inhibitor
    • lying at the boundaries between pairs of c chains within a catalytic trimer
    • three active sites
  14. o   Further examination of the ATCase-PALA complex reveals a remarkable change in quaternary structure on binding of PALA: What are these three changes?
    § the two catalytic trimers move 12 angstroms farther apart and rotate approximately 10 degrees about their common threefold axis of symmetry

    § also, the regulatory dimers rotate approximately 15 degrees to accommodate the motion

    § the enzyme expands
  15. o   In essence, ATCase has two distinct quaternary forms: one that __ and another that __
    • predominates in absence of substrate or analogs
    • predominates when substrates or analogs are bound (T and R, respectively)
  16. ·         The enzyme exists in __between the T and R state. In absence of substrate, almost all enzyme molecules are in __, with a __ and __.
    o   Binding of substrate to one active site has two effects aside from increasing likelihood that entire enzyme shifts to R state: What is the first? 

    • equilibrium 
    • T state
    • low substrate affinity and low catalytic activity

    §  Increases probability that each enzyme molecule will bind at least one substrate molecule
  17. What is the second?
    • §  Increases average number of substrate molecules bound to each enzyme; presence of additional substrate will increase the fraction of enzyme molecules in the more active R state because the position of the equilibrium depends on the number of active sites that are occupied by substrate
    • ·         The effects of substrates on allosteric enzymes= homotropic effects
  18. §  Basically, the enzyme is represented by a __mechanism
    ·         The __ for ATCase is characteristic of a mix between the T and R state curves
    o   Increasing substrate concentration favors a __; and, when switching from T to R, the curve depends on the __. It switches from T to R within a __
    • concerted 
    • sigmoidal curve
    • transition from the T-state curve to the R-state curve
    • substrate concentration
    • narrow range of substrate
  19. ·         In the T state, the __ hold the two __ sufficiently close to each other that __ and __ necessary for __ and __
    ·         Allosteric regulators modulate the T to R transition
    o   The enzyme is in the T state when bound to CTP; a binding site for this nucleotide exists in each regulatory chain in a domain that does not interact with the catalytic subunit
    §  Each active site is more than 50 angstroms from the nearest CTP-binding site
    • regulatory dimers
    • catalytic trimers
    • key loops on their surfaces collide
    • interfere with conformational adjustments
    • high-affinity substrate binding and catalysis
  20. ·         Mechanism of CTP inhibition: __
    the binding of the inhibitor CTP shifts the equilibrium toward the T state, decreasing net enzyme activity and making it more difficult for substrate binding to convert the enzyme into the R state. More substrate will be required
  21. ·         ATP is an allosteric effector in that it __ because it __
    o   The effects of nonsubstrate molecules on allosteric enzymes (like CTP and ATP) are __effects
    §  Substrates generate the __ (__effects), whereas regulators shift the__ (__ effects)
    • increases the reaction rate
    • competes with CTP for binding to regulatory sites, preventing inhibition of the enzyme
    • heterotropic 
    • sigmoidal curve
    • homotropic 
    •  KM
    • heterotropic
  22. o   Explanations for why ATP increases ATCase activity
    • §  High ATP concentration signals high concentration of purine nucleotides in the cellà lead to balance of purine and pyrimidine pools
    • §  High ATP concentration indicates that energy is available for mRNA synthesis and DNA replication and leads to the synthesis of pyrimidines needed for these processes
  23. ·         The effects of __ and __ can be modeled by changing the value of L. For the __, the value of L, which equals T/R, increases from 200 to 1250. Thus, more substrate is required to shift to R. For the ATP-saturated form, L decreases to 70
    • CTP and ATP 
    • CTP-saturated form