Supplementary Information 29 August 2013

Supplementary information 29 August 2013

1. DLAB proposal ILL Grenoble August 2013

Figure S1Dynamic Light Scattering from “native” fatted and recombinant defatted human serum albumins

Quote from “Appendix 1” of that proposal.

Light scattering at 25C in heavy water PBS from a 1mg/ml solution of “native” fatted human serum albumin (HHSA) and a 1mg/ml heavy water solution of overexpressed fully deuterated, defatted human serum albumin (DHSA).

1. Mass Spectra of DFHSA and FHSA 1mg/ml in H20 + 1% formic acid

Figure S2(a) 3782 Defatted Human Serum Albumin from ILL

Figure S2(b) 8762 Fatted Human Serum Albumin from ILL

To get these spectra trials were done to find the best conditions. Acidification with formic acid was a practical way to improve the ionisation but poor results were found for the 1mg/ml buffer solutions – as shown in Figure S3.

(a)(b)

Figure S3 Mass Spectrum of DFHSA 1mg/ml in PBS(a) No formic acid (b) with 1% formic acid

(a)(b)

Figure S4 Mass Spectrum of FHSA 1mg/ml in PBS(a) No formic acid (b) with 1% formic acid

1. DLS data H2O solutions

Figure S5 Correlation function 1µsec-1sec delay(LHS) and cumulant fitted diameter distribution function(RHS) for DFHSAfil18 at 25C

Figure S6 Correlation function 1µsec-1sec delay(LHS) and cumulant fitted diameter distribution function(RHS) for FHSAfil18 at 25C

Figure S7 Correlation function 1µsec-1sec delay(LHS) and cumulant fitted diameter distribution function(RHS) for freeze-thawed FHSAfil25 at 25C

Figure S8 Correlation function 1µsec-1sec delay(LHS) and cumulant fitted diameter distribution function(RHS) for freeze-thawed FHSAfil27 mixed at 25C

1. DLS data D2O solutions 22-7-13 to 20-8-13)

A one cumulant, fitting to the correlation function gives a weighted average for the particle radius. These radii are tabulated for each sample and the time from mixing and filtration in Table 1.

Table 1 Radii from cumultant approximation. (Values in brackets were measured using H2O values instead of D2O)

Sample / Temp / 0 days
Radius
/nm / 4 days
Radius
/nm / 16 days
Radius
/nm / Mean Radius/nm / 28 day
Radius
H2O/nm
Radius/nm / Radius/nm / Radius/nm / Radius/nm
fatted / 25°C / 5.23 / 4.04 / 3.81 / 4.36 / (4.72)
defatted / 4.14 / 4.02 / 3.97 / 4.04 / (4.71)
fatted / 4°C / 4.39 / 4.29 / 3.94 / 4.21 / (4.87)
defatted / 4.06 / 4.16 / 4.15 / 4.12 / (4.99)
fatted / -18°C / 4.07 / 4.38 / 8.37 / 5.61 / (256.28)
defatted / 4.13 / 4.43 / 5.26 / 4.61 / (55.20)
Mean r/nm / 4.33 / 4.22 / 4.91 / 4.49

As a methodological exercise the mean radii calculated in Table 1 were compared with those from the probability weighted hydrodynamic radii of the monomer peak calculated above.

Table 2 Radii from monomer peak centre of gravity.

Sample / Temp / 0 days
Radius
/nm / 4 days
Radius
/nm / 16 days
Radius
/nm / 28 days
Radius
/nm / Mean Radius/nm
Radius/nm / Radius/nm / Radius/nm / Radius/nm
fatted / 25°C / 3.49 / 3.76 / 3.95 / 4.19 / 3.84
defatted / 3.97 / 4.18 / 4.27 / 3.99 / 4.1
fatted / 4°C / 3.94 / 3.89 / 4.03 / 4.15 / 4.0
defatted / 4.12 / 3.94 / 4.06 / 3.91 / 4.0
fatted / -18°C / 4.17 / 3.96 / 3.94 / 3.86 / 3.98
defatted / 3.93 / 4.14 / 4.28 / 4.01 / 4.09
Mean
radius / 3.94 / 3.99 / 4.09 / 4.02 / 4.00

1. Small Angle X-ray Scattering (SAXS) experiments August 2013 at Bragg Institute ANSTO , Australia.

Figure S9Log-Log plot for 1.3mg/ml DHSA D2O buffer (absolute scale.

giving a spherical radius of 3.6nm.