Results tagged “detergent” from The SIBYLS Beamline

Quen Cheng in the Cooper lab has done a nice set of experiments to address the usefulness of various readily available detergents for improving the behavior proteins in SAXS experiments.

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Detergent  Category  Concentrations Shot  MW  CMC (mM)  CMC (%)  Type  Aggreg. # *  MW x N (kD) 
1-s-Nonyl-ß-D-thioglucoside 1 1x 322.4 2.9 0.093 N
C12E8 1 1x 539.1 0.1 0.006 N
C8E5 1 1x 350.5 7.1 0.249 N
CTAB 1 1x, 3x 364.5 1.0 0.036 I
DDAO 1 1X, 3x 201.4 10.4 0.209 N
DDMAB 1 0.5x, 1x, 2x 299.5 4.3 0.129 Z
FOS-Choline®-10 1 1x 323.4 13.0 0.420 Z
FOS-Choline®-12 1 1x 315.5 1.5 0.047 Z
FOS-Choline®-9 1 1x 309.4 19.0 0.588 Z
n-Decanoylsucrose 1 1x 496.6 2.5 0.124 N 55 27.31
n-Decyl-ß-D-thiomaltoside 1 1x 498.6 0.9 0.045 N 41 20.44
n-Dodecyl-N,N-dimethylglycine 1 1x 271.4 1.5 0.041 Z 76 20.63
n-Dodecyl-ß-D-maltoside 1 1x 510.6 0.2 0.009 N 100-155 65.10
n-Hexadecyl-ß-D-maltoside 1 0.5x, 1x, 2x 566.6 0.0006 0.000 N
n-Nonyl-ß-D-maltoside 1 1x 484.6 3.2 0.155 N
NP-40 1 1x, 2x 615.0 0.1 0.007
n-Tetradecyl-ß-D-maltoside 1 1x 538.6 0.01 0.001 N 120-127 66.52
n-Tridecyl-ß-D-maltoside 1 1x 524.6 0.033 0.002 N 98 51.41
n-Undecyl-ß-D-maltoside 1 1x 496.6 0.59 0.029 N 8.0-16 5.96
Zwittergent® 3-14 1 1x 363.6 0.4 0.015 Z
1-s-Heptyl-ß-D-thioglucoside 2 1x 294.4 29.0 0.854 N
1-s-Octyl-ß-D-thioglucoside 2 1x 308.4 9.0 0.278 N
C12E9 2 1x 583.1 0.1 0.005 N
C-HEGA-11 2 1x 391.5 11.5 0.450 N
C-HEGA-8 2 0.5x, 1x, 2x 293.3 180.0 5.279 N
Cymal®-1 2 1x 438.5 340.0 14.909 N
Cymal®-5 2 1x 494.5 2.4 0.119 N
Cymal®-6 2 1x 508.5 0.6 0.028 N
HECAMEG 2 1x 335.4 19.5 0.654 N
HEGA-10 2 1x 379.5 7.0 0.266 N
Heptyl-ß-D-thioglucoside 2 1x 274.3 30.0 0.823 N
IPTG 2 1x 238.3 None N 4.0-14 2.14
MEGA-9 2 1x 335.5 25.0 0.839 N
n-Decyl-ß-D-maltoside 2 1x 482.6 1.8 0.087 N
n-Nonyl-ß-D-maltoside 2 1x 468.4 6.0 0.281 N
n-Octyl-ß-D-glucoside 2 0.5x, 1x, 2x 292.4 24.5 0.716 N
Nonyl-ß-D-glucoside 2 0.5x, 1x, 2x 306.4 6.5 0.199 N
Sucrose monolaurate 2 1x 524.6 0.2 0.010 N 84 44.07
TRITON® X-100 2 0.5x, 1x, 2x 631.0 0.9 0.057 N
Zwittergent® 3-10 2 1x 307.6 40.0 1.230 Z
Zwittergent® 3-12 2 1x 335.6 4.0 0.134 Z
Zwittergent® 3-08 2 1x 279.6 330.0 9.227 Z
Anapoe® 20 3 1x 1227.5 0.059 0.007 N
Anapoe® 35 3 1x None None N
Anapoe® 56 3 1x None None N
Anapoe® 58 3 1x None None N
Anapoe® 80 3 1x 1309.7 0.012 0.002 N
Anapoe® C10E6 3 1x 427.1 0.9 0.038 N
Anapoe® C10E9 3 1x None None N
Anapoe® C12E10 3 1x None None N
Anapoe® C13E8 3 1x None None N
Anapoe® X-114 3 1x None None N
Anapoe® X-305 3 1x None None N
Anapoe® X-405 3 1x None None N
BAM 3 1x 384.5 None I
CHAPS 3 0.5x, 1x, 2x 614.9 8.0 0.492 Z
CHAPSO 3 1x 630.9 8.0 0.505 Z
C-HEGA-10 3 1x 377.5 35.0 1.321 N
C-HEGA-9 3 1x 363.5 108.0 3.926 N
Cymal®-2 3 1x 452.5 120.0 5.430 N
Cymal®-3 3 0.1x, 0.2x, 0.5x, 1x, 3x 466.5 34.5 1.609 N
Cymal®-4 3 1x 480.5 7.6 0.365 N
Deoxy BigChap 3 1x 862.1 1.4 0.121 N
FOS-Choline®-8 3 1x 295.4 102.0 3.013 Z
HEGA-8 3 1x 351.5 109.0 3.831 N
HEGA-9 3 1x 365.5 39.0 1.425 N
LDAO 3 0.5x, 1x 229.4 2.0 0.046 N
MEGA-8 3 0.1x, 1x, 3x 321.4 79.0 2.539 N
n-Hexyl-ß-D-glucoside 3 1x 264.3 250.0 6.608 N
n-Octanoylsucrose 3 1x 468.5 24.4 1.143 N 83 38.89
n-Octyl-ß-D-thiomaltoside 3 1x 470.6 9.0 0.424 N
Pluronic® F-68 3 1x ~8350 None N
Thesit® 3 1x 582.9 0.09 0.005 N

"category" - group designation based on 1xCMC scattering curve
"N" - non-ionic detergent
"I" - ionic detergent
"Z" - zwitterionic detergent
"Aggreg. #" - monomers per micelle (from CalBiochem)
"MW x N" - estimate of micelle molecular weight

Below CMC Above CMC
Group 1
(n=20)
Good for SAXS Possible with strong protein signal
Group 2
(n=22)
Good for SAXS Bad for SAXS
Group 3
(n=31)
Possible with strong protein signal Bad for SAXS

Here are the recommended concentrations of detergents that may contribute the least amount of background to SAXS studies. Omitted from this list are any detergents that would need to be used at concentrations < 0.1%, based on the above table and the detergent CMCs.

Common Detergents
0.1% CHAPS
0.2% DDMAB
0.5% OG

Group I less than 2xCMC
0.1% 1-s-Nonyl-β-D-thioglucoside
0.2% n-Decanoylsucrose
0.3% n-Nonyl-β-D-maltoside
0.4% DDAO
0.5% C8E5
0.8% FOS-Choline®-10
1.1% FOS-Choline®-9

Group II less than 1xCMC (rounded down)
0.1% Cymal®-5
0.1% Zwittergent®3-12
0.1% Nonyl-β-D-glucoside
0.2% HEGA-10
0.2% 1-s-Octyl-β-D-thioglucoside
0.2% n-Nonyl-β-D-maltoside (2)
0.4% C-HEGA-11
0.6% HECAMEG
0.8% Heptyl-β-D-thioglucoside
0.8% MEGA-9
0.8% 1-s-Heptyl-β-D-thioglucoside
1.2% Zwittergent® 3-10
5.2% C-HEGA-8
9.2% Zwittergent® 3-08
14.8% Cymal®-1

Group III less than 0.2xCMC
0.1% CHAPSO
0.2% n-Octanoylsucrose
0.3% C-HEGA-10
0.3% HEGA-9
0.3% Cymal®-3
0.5% MEGA-8
0.6% C-HEGA-9
1.0% Cymal®-2
1.2% n-Hexyl-β-D-glucoside

Detergents in SAXS

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Detergents can be extremely useful in preparing monodisperse samples for SAXS. However, they can also contribute enormous background scattering, potentially masking the desired protein signal entirely.

We recommend to not using detergents in entire protein preparation. We observed only a few cases (~0.1%) where the detergent ( bellow CMC concentration ) did not affect the protein signal. Basically, there is a high chance that detergent will make your SAXS data useless!!!!

It is crucial, therefore, to choose a detergent that will contribute the least amount of background scattering to your sample while improving sample monodispersity. The data presented here (see links at bottom of page) should be a useful starting point in choosing the right detergent for your sample. Use a detergent only in the case when the protein preparation without detergent was not successful

How we looked for good detergents

We searched for SAXS-compatible detergents by collecting data on 73 detergents in PBS. By subtracting the PBS signal from the detergent signal, we can observe the scattering contribution of detergent alone. We first collected data at 1xCMC (critical micelle concentration) to compare the scattering effects of different detergent types. Next, we investigated the effects of concentration relative to the CMC, collecting data ranging from 0.1xCMC to 3xCMC on a subset of the 73 detergents.

Observed Trends

Scattering signal is highly concentration-dependent (below CMC is much better than above CMC), but not all detergents are created equal. Based on the 1xCMC scattering curves, we derived three groups of detergents with scattering properties ranging from essentially no signal (group 1) to large protein-like signal (group 3). The following table provides a rough guide of detergent SAXS-compatibility:

Below CMC Above CMC
Group 1
(n=20)
Good for SAXS Possible with strong protein signal
Group 2
(n=22)
Good for SAXS Bad for SAXS
Group 3
(n=31)
Possible with strong protein signal Bad for SAXS

Caveats

  • 1xCMC may not be a very reliable data point
    • Sensitive to pipetting error
    • CMC fluctuates with temperature, pH, ionic strength
  • Majority of data from single experiment
  • Unclear why detergents scatter differently; no obvious trends in MW, micelle size, % concentration, etc.
  • Detergents alone may behave differently than detergents in complex with protein
  • Some level of subjectivity in "group" designations
  • Doesn't address membrane protein applications

What the data provides, however, is a starting point for finding a detergent that improves the monodispersity of your sample without dominating the SAXS signal.

Data

The following table include links to scattering curves of each concentration of detergent we analyzed, with Glucose Isomerase plotted as a reference protein curve.

A sortable Table of detergents.

List of best-bet detergents

ALS Ring Status

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Funding

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