NMR lecture dynamics - NMR Spectroscopy - Protein dynamics. Spectral density function Rotational...

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Transcript of NMR lecture dynamics - NMR Spectroscopy - Protein dynamics. Spectral density function Rotational...

  • NMR Spectroscopy Protein dynamics

  • NMR Spectroscopy - Protein dynamics Chemical exchange

    A B k

    k

    N

    CH 3

    CH 3 O

    N k

    k N

    CH 3

    CH 3

    O

    N

    Average time between jumps is !=1/k τ=1/k

  • NMR Spectroscopy - Protein dynamics Chemical exchange

    k vs νA – νB

  • NMR Spectroscopy - Protein dynamics Chemical exchange

  • NMR Spectroscopy - Protein dynamics Chemical exchange

  • NMR Spectroscopy - Protein dynamics Chemical exchange

  • NMR Spectroscopy - Protein dynamics NMR vs UV time scales

  • NMR Spectroscopy - Protein dynamics Chemical Exchange - J coupling

  • Relaxation mechanisms

    Longitudinal relaxation requires a time-dependent magnetic field fluctuating at the Larmor frequency

    The time-dependence originates in the motions of the molecule (vibration, rotation, diffusion etc)

    Molecules in solution “tumble”. This “tumbling” can be characterized by a rotational correlation time τc

    τc is the time needed for the rms deflection of the molecules to be ~ 1 radian (60°)

    NMR Spectroscopy - Protein dynamics

  • Spectral density function

    Rotational diffusion in solution occurs at a range of frequencies

    1/τc ~ rms rotational frequency (radians s-1)

    The probability function of finding motions at a given angular frequency ω can be described by the spectral density function J(ω)

    NMR Spectroscopy - Protein dynamics

  • Spectral density function

    Frequency distribution of the fluctuating magnetic fields

    NMR Spectroscopy - Protein dynamics

  • Spectral density function - Relaxation rates

    NMR Spectroscopy - Protein dynamics

  • Relaxation rates

    NMR Spectroscopy - Protein dynamics

  • NMR Spectroscopy - Protein dynamics

  • NMR Spectroscopy - Protein dynamics Spectral density mapping

    Reduced spectral density mapping

    J(ωH)~ J(ωH – ωN)~ J(ωH + ωN)~ J()

  • NMR Spectroscopy - Protein dynamics Spectral density mapping

  • NMR Spectroscopy - Protein dynamics Model free approach

    where 1/τ1’=
1/τC– 1/τ1

    where S2, order parameter

  • NMR Spectroscopy - Protein dynamics Model free approach

  • NMR Spectroscopy - Protein dynamics Slow vs Fast motions

    ps-ns

    μs-ms

  • NMR Spectroscopy - Protein dynamics Hydrogen/Deuterium (H/D) exchange

    kop kex kcl

  • NMR Spectroscopy - Protein dynamics Hydrogen/Deuterium (H/D) exchange

  • NMR Spectroscopy - Protein dynamics Hydrogen/Deuterium (H/D) exchange

    kop kch kcl

    kobs=kop kch/(kop + kcl + kch)

    EX2: kcl>>kch kobs=kop kch/(kcl) = Kop kch

    Kop is referred to as the protection factor, P

    ΔGop = –RTlnKop

  • NMR Spectroscopy - Protein dynamics

    Significant heterogeneity in the magnitude of the protection factors

    Hydrogen/Deuterium (H/D) exchange

    A large number of states define the native-state ensemble

  • NMR Spectroscopy - Protein dynamics Hydrogen/Deuterium (H/D) exchange

    kop kch kcl

    kobs=kop kch/(kop + kcl + kch)

    EX1: kcl

  • NMR Spectroscopy - Protein dynamics Hydrogen/Deuterium (H/D) exchange

    kop

    kch (s -1)

    N-H (closed)

    kcl

    N-H (open) kch

    D2O

    [kobs=kop]

    [kobs=(kop/kcl) kch)]

  • NMR Spectroscopy - Protein dynamics Hydrogen/Deuterium (H/D) exchange

    6

    12

    ΔΔ G

    ( kc

    al m

    ol -1 )

  • NMR Spectroscopy - Protein dynamics Hydrogen/Deuterium (H/D) exchange

  • NMR Spectroscopy - Protein dynamics Hydrogen/Deuterium (H/D) exchange

  • NMR Spectroscopy - Protein dynamics Hydrogen/Deuterium (H/D) exchange

  • NMR Spectroscopy - Protein dynamics An illustrative system

  • NMR Spectroscopy - Protein dynamics Chemical shift mapping

  • NMR Spectroscopy - Protein dynamics Slow dynamics (μs-ms)

  • NMR Spectroscopy - Protein dynamics Slow dynamics (μs-ms)

  • NMR Spectroscopy - Protein dynamics Fast dynamics (ps-ms)

  • NMR Spectroscopy - Protein dynamics Changes in order parameters, S2

  • NMR Spectroscopy - Protein dynamics Changes in order parameters, S2

  • NMR Spectroscopy - Protein dynamics From dynamics to entropy

  • NMR Spectroscopy - Protein dynamics From dynamics to entropy

  • NMR Spectroscopy - Protein dynamics Thermodynamics of ligand binding

  • NMR Spectroscopy - Protein dynamics Dynamics may drive ligand binding