Cooperatively and Allosteric properties

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Cooperatively and Allosteric properties

• Or
• Making Things Easy and Regulation

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Cooperatively and Allosteric properties

• Often there is more than one binding site in a
  protein we can use binding studies to look at
  several properties.
• Number of sites
• Tightness of binding at sites
• are the sites independent or do they interact.
• Caveat Assumptions in mathematical models dont
  always match reality.

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Definitions

• Allosteric Change in the shape and activity of a
  protein by the binding of a ligand to a site
  other than the chemically active site.
• Cooperativety Each step in a multi-step process
  changes the likely hood of the next step
  occurring
• positive enhances
• negative inhibits

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Cooperatively and Allosteric properties

• For sites that show a normal hyperbolic binding
  curves ligand vs Bound curve Scatchard analysis
  is a valuable tool.
• The assumption is that binding at each site is
  independent of one another.
• Sites may have identical Ka or differering Ka
• We plot PA vs PA/L
• The result suggest the number of sites in the
  molecule.

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Cooperatively and Allosteric properties
Deviations form linearity tell us that sites are
not equivalent Differing binding constants or
cooperatively
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Cooperative effects in Hemoglobin

• The initial binding of O2 is difficult.
• Binding of the second and third are progressively
  easier
• The fourth O2 is energetically free

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Cooperative effects in Hemoglobin

• If we see a non hyperbolic binding curve then we
  are interested in pursuing a Hill coefficient to
  help explain what is happening.

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Cooperative effects in Hemoglobin

• What is assumed is that there are no
  intermediates a protein is fully occupied or it
  is completely unoccupied

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Cooperative effects in Hemoglobin

• Binding is never totally Cooperative so we
  typically see plots with three portions the ends
  of the plots have slopes of 1 or non cooperative
  binding and the middle has a slope other than 1

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Cooperative effects in Hemoglobin

• Slope less than one indicate negitive
  coopertivaty.
• Hemoglobin has a slope greater than one binding
  is positively cooperative

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Heterotropic Allosteric Effector in Hemoglobin

• BPG binds preferentially to the tense (lower
  binding form) of hemoglobin (see molecular
  models).
• It shifts the binding curve to the right, higher
  concentrations of oxygen are required to fully
  saturate hemoglobin.
• Oxygen delivery at low oxygen pressure (high
  altitude) is enhanced by increasing the amount of
  diphosphoglycerate in the red blood cells.

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Cooperatively and Allosteric properties

• Sequential model of binding
• Ligands may only bind to the relaxed form
• Each binding converts the protein domain or
  subunit from a tense form to a relaxed form
• each binding also forces adjacent units towards
  the relaxed form.

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Cooperatively and Allosteric properties

• concerted model of binding
• Protein in equilibrium between tense and relaxed
  form
• Ligand may bind to tense or relaxed form
• ligand binding stabilizes relaxed form
• each additional ligand provides additional
  stabilty for relaxed form.