Impact of protein

1
Impact of protein detergent interactions on NMR
structure determination
Linda Columbus Department of Chemistry University
of Virginia
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Membrane Proteins
BtuB
LamB
OmpX
OM
KcsA
SecYEG
BtuCD
CM
lt 100 polytopic membrane protein structures
deposited in the pdb compared to 37,000 soluble
protein structures 55 of bacterial membrane
proteins have unknown functions 60 of drugs on
the market are targeted to membrane proteins
3
NMR membrane protein structures
OmpG
PagP
dodecylphosphocholine octyl
glucoside
dodecylphosphocholine
OmpA
OmpX
1, 2-dihexanoyl-sn-glycero-phosphocholine
dodecylphosphocholine
4
The role of detergents in membrane protein
structural and functional studies
5
The role of detergents in membrane protein
structural and functional studies
6
The role of detergents in membrane protein
structural and functional studies
7
The role of detergents in membrane protein
structural and functional studies
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Properties of detergent micelles

• Critical micelle concentration (cmc)
• Concentration of monomer in equilibrium with
  micelle
• Aggregation number
• Number of monomers in micelle
• Shapes and sizes prolate, oblate, cylinder,
  etc...

9
Model System TM0026
Preliminary structural model
15N (ppm)
C
N
1H (ppm)
10
Detergents influence the completeness of NMR
observations and tertiary interactions
DM

• What process in DDM and FC-10 is contributing to
  line broadening?

DDM
2. Why do DM and FC-12 PDCs have reduced line
broadening?
FC-10
FC-12
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• What happens in FC-10 and DDM that contributes to
  line broadening?

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SDSL indicates the helical interaction is
disrupted in FC-10
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Why do DM and FC-12 give rise to quality NMR
spectra?
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Geometry of micelle influences the completeness
of NMR observations and tertiary interactions
Detergent (abbreviation) Ag L (Å) Shperoid rHC (Å) VHC (nm3) Ionic property
n-decyl-?-D-maltoside (DM) 80-90 34 oblate 12, 23 26 Non-ionic
n-dodecyl-?-D-maltoside (DDM) 135-145 39 oblate 14, 29 49 Non-ionic
n-decylphosphocholine (FC-10) 45-53 28 prolate 21, 13 15 zwitterionic
n-dodecylphosphocholine (FC-12) 65-80 34 prolate 26, 16 27 zwitterionic
Detergent (abbreviation) Ag L (Å) Shperoid rHC (Å) VHC (nm3) Ionic property
n-decyl-?-D-maltoside (DM) 80-90 34 oblate 12, 23 26 Non-ionic
n-dodecyl-?-D-maltoside(DDM) 135-145 39 oblate 14, 29 49 Non-ionic
n-decylphosphocholine (FC-10) 45-53 28 prolate 21, 13 15 zwitterionic
n-dodecylphosphocholine (FC-12) 65-80 34 prolate 26, 16 27 zwitterionic
DDM/FC-10 (molar ratio 47) 77 (30/47) 34 oblate 14, 21 26 zwitterionic
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Hot off the spectrometer
Detergent (abbreviation) N Ionic property
n-octyl-?-D-glucopyranoside (OG) 75-100 Non-ionic
n-nonyl-?-D-glucopyranoside (NG) 160-180 Non-ionic
n-decyl-?-D-glucopyranoside (DG) ND Non-ionic
n-decyl-?-D-maltoside (DM) 70-75 Non-ionic
n-dodecyl-?-D-maltoside (DoDM) 130-140 Non-ionic
1,2-dihexanoyl-sn-glycero-phosphocholine (DHPC) 35-40 zwitterionic
3-(3-cholamidopropyl)-dimethylammonio-1-propane sulfonate (CHAPS) 14-18 zwitterionic
Lauryldimethylyamine oxide (LDAO) NC zwitterionic
n-decylphosphocholine (FC-10) 35-45 zwitterionic
n-dodecylphosphocholine (FC-12) 65-70 zwitterionic
1-palmitoyl-2-hydroxy-sn-glycero-3-phospho-rac-(1-glycerol) (LPPG) 100-105 ionic
Dan Fox and Kalyani Jambunathan
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Unique structure is stabilized by a hydrophobic
match to the detergent shape and size
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Acknowledgements
Small Angle X-ray Scattering
Jan Lipfert Sebastian Doniach

• Scott Lesley
• Gerard Kroon
• Bernhard Geierstanger

Kalyani Bill Brett
Dan Jambunathan Peairs
Kroncke Fox
Iza Thien Chris
Justin Bielnicka
Nguyen Reyes Kim
NRSA 1F32GM068286 UVA Start-up funds