Collision Cross Sections of Proteins and their Complexes: a ...
Collision Cross Sections of Proteins and their Complexes: a Calibration Framework and Database for Gas-Phase Structural Biology
Matthew F. Bush,a Zoe Hall,a Kevin Giles,b John Hoyes,b Carol V. Robinson,a* Brandon T. Ruotoloc*
Supporting Information
a
b
Department of Chemistry, Physical and Theoretical Chemistry Laboratory, University of Oxford, South Parks Road, Oxford OX1 3QZ, United Kingdom
Waters Corporation, Floats Road, Wythenshawe , Manchester, M23 9LZ, United Kingdom c
Department of Chemistry, University of Michigan, Ann Arbor, MI, 48109, United States
*Address communication to: [email protected], [email protected]
S1
Average Effective Densities The effective protein complex radius depends on the average He measured for all observed charge states (He) and the contribution of He to the impact parameters (ݎு , approximated as 1 Å): ݎ ൌ ට
ۃஐಹ ۄ గ
െ ݎு
(1)
The effective protein complex volume is calculated using: ସ
ଷ ܸ ൌ ߨݎ ଷ
(2)
Finally, effective densities, Deff, are then determined using the molecular weight (MW) of the ion and Avogadro’s number (N0): ܦ ൌ
ெௐ
ଵ
ேబ
S2
(3)
Table S1. Sample Sources and Preparation Protocols Sample
Supplier
Item #
Protocol
Gly-Arg-Gly-Asp-Ser
Sigma
G4391
DN
Ser-Asp-Gly-Arg-Gly
Sigma
S3771
DN
additional peptides
Waters
186002337
DN
ubiquitin
Sigma
U6253
DN
cytochrome c
Fluka
30396
myoglobin
Sigma
M1882
DN
-lactoglobulin A
Sigma
L7880
Buffer
transthyretin
Sigma
P1742
Buffer, BS
avidin
Sigma
A9275
Buffer
bovine serum albumin
Sigma
P7656
Buffer
concanavalin A
Sigma
C2010
Buffer, BS
serum amyloid P
CalBioChem
565190
Buffer, BS
alcohol dehydrogenase
Sigma
A7011
Buffer, BS
pyruvate kinase
Sigma
P9136
Buffer, BS twice
glutamate dehydogenase
Sigma
G7882
Buffer, BS twice
GroEL
Sigma
C7688
Refolded,1 Buffer, BS twice
DN Buffer
DN = Samples prepared in water/methanol/acetic acid (49/49/2) solutions. Buffer = Samples prepared in 200 mM aqueous ammonium acetate solutions, then stored in 10 – 20 L aliquots at –20 °C. Ions from buffered solutions are characterized using very gentle instrumental conditions, near the threshold for ion transmission. Ions characterized under these conditions will be referred to as “native like”. BS = On day of analysis, sample buffer exchanged using a Micro Bio-Spin 6 column (Bio-Rad, Hercules, CA) that has been equilibrated with 200 mM aqueous ammonium acetate.1
S3
Table S2. Collision Cross Section () for Denatured Ions m / Da Z GRGDS
490
SDGRG
490
Angiotensin fragment 1-7 RASG-1 Angiotensin II Bradykinin Angiotensin I Renin substate Enolase T35 Enolase T37
ubiquitin bovine erythrocytes
898 1 000 1 046 1 060 1 296 1 758 1 872 2 827
8k
cytochrome c equine heart
12 k
apo myoglobin equine heart
17 k
1 2 1 2 2 2 2 2 3 3 3 3 7 8 9 10 11 12 13 13 14 15 16 17 18 19 20 15 16 17 18 19 20 21 22 23 24 25 26
S4
He / nm2 1.32 1.39 1.30 1.42 2.26 2.25 2.45 2.37 3.28 3.80 3.80 4.65 16.7 17.3 18.0 18.9 19.8 25.2 26.0 26.7 27.4 28.0 28.7 29.2 35.2 36.0 36.8 37.5 38.2 38.7 39.2 39.6 40.0
N2 / nm2 2.06 2.56 2.04 2.59 3.34 3.31 3.35 3.44 4.74 5.22 5.19 19.1 19.9 20.9 22.0 23.4 24.8 26.0 30.8 32.0 33.3 34.5 36.0 36.7 37.9 40.6 41.8 43.1 44.4 45.7 47.0 48.2 49.2 50.1 50.9 -
Table S3. Collision Cross Sections () for Native-Like Ions n
m / Da
1
12 k
1
18 k
2
37 k
transthyretin human plasma
4
56 k
avidin egg white
4
64 k
serum albumin bovine
1
69 k
concanavalin A Canavalia ensiformis
4
103 k
serum amyloid P human serum
5
125 k
alcohol dehydogenase Saccharomyces cerevisiae
4
143 k
pyruvate kinase rabbit heart
4
237 k
serum amyloid P human serum
10
250 k
cytochrome c equine heart -lactoglobulin bovine milk
S5
z 6 7 7 8 9 11 12 13 14 15 16 15 16 17 18 14 15 16 17 19 20 21 22 23 22 23 24 25 26 23 24 25 26 30 31 32 33 34 35 31 32 33
He / nm2 12.4 12.8 16.6 16.9 17.8 28.5 29.0 29.6 34.1 34.0 33.8 36.4 36.4 36.4 36.4 40.9 41.0 40.6 40.4 55.5 55.5 54.8 54.5 70.3 69.7 69.3 68.6 68.3 69.4 69.4 68.3 67.2 103 103 103 102 102 100 104 105 106
N2 / nm2 14.9 15.9 19.5 20.3 32.3 33.1 34.3 38.4 38.5 38.8 41.5 41.5 41.6 44.9 44.9 44.7 44.9 60.6 60.8 60.9 60.5 76.3 76.0 74.6 73.1 72.8 74.2 74.5 74.4 75.0 111 111 110 110 110 111 112 112
glutamate dehydrogenase bovine liver
GroEL Escherichia coli
6
336 k
14
801 k
S6
34 35 37 38 39 40 41 42 43 65 66 67 68 69 70 71 72
105 107 128 128 128 128 128 128 209 209 207 207 206 207
111 134 134 134 134 135 218 220 220 219 219 218 219 -
Figure S1. Absolute collision cross sections are determined directly from the slope of drift time versus reciprocal drift-voltage plots. The intercepts with the drift time axis correspond to t0. Drift times for the observed charge states of denatured peptide (DN Peptides), denatured cytochrome c (DN Cyt c), native-like transthyretin tetramer (TTR), native-like concanavalin A tetramer (Conc A), and native-like glutamate denhydrogenase hexamer (Glu Dhn) ions were measured at 10 drift voltages ranging from 60–200 V in 2.0 Torr of nitrogen gas. R2 values for the best fit lines are > 0.9997 and the uncertainties for the slopes are < 0.6 %.
S7
Figure S2. Calibrating travelling-wave drift times using all denatured protein and native-like protein complex ions (A). Travelling-wave drift times were measured using 0.4 Torr N2, a wave velocity of 300 m/s, and wave heights (WH) of 8 V (red squares), 9 V (green triangles), or 10 V (blue circles). Expansion of data for denatured protein ions (B). Correlation coefficients in B are for the correlation between data for the denatured protein ions and the calibration plot obtained using only data for native-like protein complex ions
S8
References 1.
Hernandez, H.; Robinson, C. V. Nature Prot. 2007, 2, 715-726.
S9
Matthew F. Bush,a Zoe Hall,a Kevin Giles,b John Hoyes,b Carol V. Robinson,a* Brandon T. Ruotoloc*
Supporting Information
a
b
Department of Chemistry, Physical and Theoretical Chemistry Laboratory, University of Oxford, South Parks Road, Oxford OX1 3QZ, United Kingdom
Waters Corporation, Floats Road, Wythenshawe , Manchester, M23 9LZ, United Kingdom c
Department of Chemistry, University of Michigan, Ann Arbor, MI, 48109, United States
*Address communication to: [email protected], [email protected]
S1
Average Effective Densities The effective protein complex radius depends on the average He measured for all observed charge states (He) and the contribution of He to the impact parameters (ݎு , approximated as 1 Å): ݎ ൌ ට
ۃஐಹ ۄ గ
െ ݎு
(1)
The effective protein complex volume is calculated using: ସ
ଷ ܸ ൌ ߨݎ ଷ
(2)
Finally, effective densities, Deff, are then determined using the molecular weight (MW) of the ion and Avogadro’s number (N0): ܦ ൌ
ெௐ
ଵ
ேబ
S2
(3)
Table S1. Sample Sources and Preparation Protocols Sample
Supplier
Item #
Protocol
Gly-Arg-Gly-Asp-Ser
Sigma
G4391
DN
Ser-Asp-Gly-Arg-Gly
Sigma
S3771
DN
additional peptides
Waters
186002337
DN
ubiquitin
Sigma
U6253
DN
cytochrome c
Fluka
30396
myoglobin
Sigma
M1882
DN
-lactoglobulin A
Sigma
L7880
Buffer
transthyretin
Sigma
P1742
Buffer, BS
avidin
Sigma
A9275
Buffer
bovine serum albumin
Sigma
P7656
Buffer
concanavalin A
Sigma
C2010
Buffer, BS
serum amyloid P
CalBioChem
565190
Buffer, BS
alcohol dehydrogenase
Sigma
A7011
Buffer, BS
pyruvate kinase
Sigma
P9136
Buffer, BS twice
glutamate dehydogenase
Sigma
G7882
Buffer, BS twice
GroEL
Sigma
C7688
Refolded,1 Buffer, BS twice
DN Buffer
DN = Samples prepared in water/methanol/acetic acid (49/49/2) solutions. Buffer = Samples prepared in 200 mM aqueous ammonium acetate solutions, then stored in 10 – 20 L aliquots at –20 °C. Ions from buffered solutions are characterized using very gentle instrumental conditions, near the threshold for ion transmission. Ions characterized under these conditions will be referred to as “native like”. BS = On day of analysis, sample buffer exchanged using a Micro Bio-Spin 6 column (Bio-Rad, Hercules, CA) that has been equilibrated with 200 mM aqueous ammonium acetate.1
S3
Table S2. Collision Cross Section () for Denatured Ions m / Da Z GRGDS
490
SDGRG
490
Angiotensin fragment 1-7 RASG-1 Angiotensin II Bradykinin Angiotensin I Renin substate Enolase T35 Enolase T37
ubiquitin bovine erythrocytes
898 1 000 1 046 1 060 1 296 1 758 1 872 2 827
8k
cytochrome c equine heart
12 k
apo myoglobin equine heart
17 k
1 2 1 2 2 2 2 2 3 3 3 3 7 8 9 10 11 12 13 13 14 15 16 17 18 19 20 15 16 17 18 19 20 21 22 23 24 25 26
S4
He / nm2 1.32 1.39 1.30 1.42 2.26 2.25 2.45 2.37 3.28 3.80 3.80 4.65 16.7 17.3 18.0 18.9 19.8 25.2 26.0 26.7 27.4 28.0 28.7 29.2 35.2 36.0 36.8 37.5 38.2 38.7 39.2 39.6 40.0
N2 / nm2 2.06 2.56 2.04 2.59 3.34 3.31 3.35 3.44 4.74 5.22 5.19 19.1 19.9 20.9 22.0 23.4 24.8 26.0 30.8 32.0 33.3 34.5 36.0 36.7 37.9 40.6 41.8 43.1 44.4 45.7 47.0 48.2 49.2 50.1 50.9 -
Table S3. Collision Cross Sections () for Native-Like Ions n
m / Da
1
12 k
1
18 k
2
37 k
transthyretin human plasma
4
56 k
avidin egg white
4
64 k
serum albumin bovine
1
69 k
concanavalin A Canavalia ensiformis
4
103 k
serum amyloid P human serum
5
125 k
alcohol dehydogenase Saccharomyces cerevisiae
4
143 k
pyruvate kinase rabbit heart
4
237 k
serum amyloid P human serum
10
250 k
cytochrome c equine heart -lactoglobulin bovine milk
S5
z 6 7 7 8 9 11 12 13 14 15 16 15 16 17 18 14 15 16 17 19 20 21 22 23 22 23 24 25 26 23 24 25 26 30 31 32 33 34 35 31 32 33
He / nm2 12.4 12.8 16.6 16.9 17.8 28.5 29.0 29.6 34.1 34.0 33.8 36.4 36.4 36.4 36.4 40.9 41.0 40.6 40.4 55.5 55.5 54.8 54.5 70.3 69.7 69.3 68.6 68.3 69.4 69.4 68.3 67.2 103 103 103 102 102 100 104 105 106
N2 / nm2 14.9 15.9 19.5 20.3 32.3 33.1 34.3 38.4 38.5 38.8 41.5 41.5 41.6 44.9 44.9 44.7 44.9 60.6 60.8 60.9 60.5 76.3 76.0 74.6 73.1 72.8 74.2 74.5 74.4 75.0 111 111 110 110 110 111 112 112
glutamate dehydrogenase bovine liver
GroEL Escherichia coli
6
336 k
14
801 k
S6
34 35 37 38 39 40 41 42 43 65 66 67 68 69 70 71 72
105 107 128 128 128 128 128 128 209 209 207 207 206 207
111 134 134 134 134 135 218 220 220 219 219 218 219 -
Figure S1. Absolute collision cross sections are determined directly from the slope of drift time versus reciprocal drift-voltage plots. The intercepts with the drift time axis correspond to t0. Drift times for the observed charge states of denatured peptide (DN Peptides), denatured cytochrome c (DN Cyt c), native-like transthyretin tetramer (TTR), native-like concanavalin A tetramer (Conc A), and native-like glutamate denhydrogenase hexamer (Glu Dhn) ions were measured at 10 drift voltages ranging from 60–200 V in 2.0 Torr of nitrogen gas. R2 values for the best fit lines are > 0.9997 and the uncertainties for the slopes are < 0.6 %.
S7
Figure S2. Calibrating travelling-wave drift times using all denatured protein and native-like protein complex ions (A). Travelling-wave drift times were measured using 0.4 Torr N2, a wave velocity of 300 m/s, and wave heights (WH) of 8 V (red squares), 9 V (green triangles), or 10 V (blue circles). Expansion of data for denatured protein ions (B). Correlation coefficients in B are for the correlation between data for the denatured protein ions and the calibration plot obtained using only data for native-like protein complex ions
S8
References 1.
Hernandez, H.; Robinson, C. V. Nature Prot. 2007, 2, 715-726.
S9
