[RASMB] another vbar question

Rocco Mattia mattia.rocco at hsanmartino.it
Thu Sep 28 07:00:00 PDT 2017


Hello Holger,
 
nice work you're doing with vbar... perhaps it will lead to some correction factors to be applied to calculated vbars as a function of salt type/molarity...? ;-)
 
Not surprising that vbar varies with buffer conditions. It's an operational quantity derived from the molar volume of compounds in solution, and that volume can change because of ion effects, solvent preferential binding, etc. The way it's calculated, it's based on measured values for particular groups under standad conditions (i.e. water at 20 °C, or sometimes at 4 °C). Clearly, if the molar volumes change as a function of the solution conditions, so the vbar will change... A notorious case is DNA, for which measured values of vbar are strongly dependent, say, on Mg content....
 
Although for most standard uses, such as the calculation of MW from AUC or SAXS, and under "normal" conditions, the errors in vbar won't affect too much the answer(s) sought (It's a monomer or a dimer....?), if one really wants to be precise, the vbar should be measured.... apart from the DM500, differential sedimentation in H2O(16) vs H2O(18) should provide the answer with much less material (but of course H2O(18) is quite expensive, and dialysis is required....)
 
Cheers - Mattia

________________________________

From: RASMB on behalf of HGSR (Holger Martin Strauss)
Sent: Thu 9/28/2017 3:48 PM
To: rasmb at list.rasmb.org
Subject: [RASMB] another vbar question


Hello yall,
 
Wow, vbar trending?!
 
I actually have a question regarding the salt-dependency of the vbar. We are working with peptides modified with some organic linkers and a diacid (between C4-C50, to give a range). We measure the droh/dc (DMA5000) at different concentrations and calculate the vbar. Controls OK, concentration accurate and measurements reproducible, buffer dialysed. Salt-concentration is varied (NaCl, 5-1500 mM-ish). Turns out the vbar is considerably different at high salt. (And so is the self-association, btw.) No detergents or denaturants, just some additional phosphate for buffering, besides the NaCl.
 

	
*	How can this be rationalised structurally? 
*	Any tests when droh/dc is more appropriate for Mw /sigma than vbar? 
*	Other examples in the literature for peptides? 
*	Any general comments/anectodes/remarks? 

 
Cheers, 
 
Holger
 


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