Hi

I'd like to use SASREFmx to analyse samples with polydispersity. However, I'm struggling without a manual. I'd assumed it would run in a similar manner to SASREF but I'm confused as to what goes into the .con file requested at the beginning- is this asking for contact or scattering information?

Many thanks

Mike

## sasrefmx

### Re: sasrefmx

The Atsas-style documentation is currently missing for Sasrefmx and will become available in the next Atsas release

The easiest way at present is to run SASREFMX via Atsas-online

by selecting the option "SAXS only, polydisperse system"

If needed, one may re-use the con files also for the local runs of Sasrefmx.

Finally, the Sasrefmx manual can be provided at the moment in a form of a text file.

The easiest way at present is to run SASREFMX via Atsas-online

by selecting the option "SAXS only, polydisperse system"

If needed, one may re-use the con files also for the local runs of Sasrefmx.

Finally, the Sasrefmx manual can be provided at the moment in a form of a text file.

### Re: sasrefmx

Hi

Many thanks for the quick reply. I'm running SASERFMX online, and am trying to understand how the curve sub-unit correspondence is filled in.

The online instruction is:

"For dissociation products use 0.0 if a subunit belongs to a subconstruct or (for the free subunit) its molar ratio in the mixture"

I have two proteins forming a dimer (A+B=AB), which then oligomerises to 2(AB) or possibly 3(AB). If I want to account for polydispersity caused by free A, do I represent a 10% molar ratio of the free A protein with 0.1 in the 'dissociation products' field, and 0.9 in the 'scattering data.dat' field?

I'd love to have a look at the manual if that can be emailed

Thanks for your help

Mike

mher065@aucklanduni.ac.nz

Many thanks for the quick reply. I'm running SASERFMX online, and am trying to understand how the curve sub-unit correspondence is filled in.

The online instruction is:

"For dissociation products use 0.0 if a subunit belongs to a subconstruct or (for the free subunit) its molar ratio in the mixture"

I have two proteins forming a dimer (A+B=AB), which then oligomerises to 2(AB) or possibly 3(AB). If I want to account for polydispersity caused by free A, do I represent a 10% molar ratio of the free A protein with 0.1 in the 'dissociation products' field, and 0.9 in the 'scattering data.dat' field?

I'd love to have a look at the manual if that can be emailed

Thanks for your help

Mike

mher065@aucklanduni.ac.nz

### Re: sasrefmx

For Sasrefmx, one needs to know the oligomeric state of the largest assembly,

so if you are unsure if it is 2(AB) or 3(AB), you need to consider these cases separately.

The situation when there are both 2(AB), 3(AB) and free A in solution cannot be handled (at least at the moment).

Do you expect symmetric AB arrangement, e.g. P2 for 2(AB) or P3 for 3(AB) ?

Can free A multimerize as well?

The "molar ratio in the mixture" means the ratio in the original mixture (before binding).

It is used when you mix let say two moles of A with one mole of B and let them form a partial complex,

so that there will be (2-x) moles of free A, (1-x) moles of free B and x moles of the complex.

In this case one enters 2 for A and 1 for B in the table.

If I understood you right, in your case there is simply an excess of A,

so that you use 0.0 for A and -1.0 for B.

Please see corresponding screenshots of SasrefMx for the case of dimeric complex plus excess of monomeric A:

so if you are unsure if it is 2(AB) or 3(AB), you need to consider these cases separately.

The situation when there are both 2(AB), 3(AB) and free A in solution cannot be handled (at least at the moment).

Do you expect symmetric AB arrangement, e.g. P2 for 2(AB) or P3 for 3(AB) ?

Can free A multimerize as well?

The "molar ratio in the mixture" means the ratio in the original mixture (before binding).

It is used when you mix let say two moles of A with one mole of B and let them form a partial complex,

so that there will be (2-x) moles of free A, (1-x) moles of free B and x moles of the complex.

In this case one enters 2 for A and 1 for B in the table.

If I understood you right, in your case there is simply an excess of A,

so that you use 0.0 for A and -1.0 for B.

Please see corresponding screenshots of SasrefMx for the case of dimeric complex plus excess of monomeric A:

### Re: sasrefmx

Hi,

out of curiosity I would like to understand how the linear combination in SasrefMX is performed, since the scattering curves of the components and the coefficients aren't included in the atsasonline version of the program.

I mean:

I have the experimental scattering pattern of an A2B2 complex, and I tried to fit it admitting partial dissociation.

The result is "associated fraction 0.77"

So, I guess the calculated intensity fitting the curve is Icalc=ScaleFactor*(f1*IA2B2 + f2*IA + f3*IB) where f1, f2 and f3 are volume fractions and intensities are calculated using Crysol.

So if f1=0.77, then should f2 and f3 be calculated taking into account that subunits A and B have different volumes VA and VB, right?

A and B molecules should be present in the same number since they come from A2B2 dissociating in 2*(A+B) but since they occupy different volumes I think f2 and f3 are obtained considering that f2:f3=VA/VB.

Could anybody tell me if I am right or not?

out of curiosity I would like to understand how the linear combination in SasrefMX is performed, since the scattering curves of the components and the coefficients aren't included in the atsasonline version of the program.

I mean:

I have the experimental scattering pattern of an A2B2 complex, and I tried to fit it admitting partial dissociation.

The result is "associated fraction 0.77"

So, I guess the calculated intensity fitting the curve is Icalc=ScaleFactor*(f1*IA2B2 + f2*IA + f3*IB) where f1, f2 and f3 are volume fractions and intensities are calculated using Crysol.

So if f1=0.77, then should f2 and f3 be calculated taking into account that subunits A and B have different volumes VA and VB, right?

A and B molecules should be present in the same number since they come from A2B2 dissociating in 2*(A+B) but since they occupy different volumes I think f2 and f3 are obtained considering that f2:f3=VA/VB.

Could anybody tell me if I am right or not?

### Re: sasrefmx

Yes, you are right,

if you used equimolar ratio (i.e. specified 1 and 1 in the last row of the table) for the dissociation products,

then the ratio between their volume fractions is equal to Va/VB

if you used equimolar ratio (i.e. specified 1 and 1 in the last row of the table) for the dissociation products,

then the ratio between their volume fractions is equal to Va/VB

### Re: sasrefmx

Yes I actually wrote in the second line:SaxsMax wrote:Yes, you are right,

if you used equimolar ratio (i.e. specified 1 and 1 in the last row of the table) for the dissociation products,

then the ratio between their volume fractions is equal to Va/VB

a1.pdb b1.pdb a2.pdb b2.pdb

0.1 0.1 0.1 0.1

thinking that those number were molar fractions but I think that equimolarity is satisfied as well

Thank you