[RASMB] another vbar question

Fri Sep 29 01:59:00 PDT 2017

Hej Arne and all,

We’re thinking along similar lines here (and thanks for sharing that much here. Question is, where’s the acyl chain for the solvated particle?); from my experience, the vbar/molar increments for peptides are basically useless, which is why we measure them. Distinguishing a 1-2 equ. Might be doable from sigma alone, but not the difference between a 1-2-3 and 2-4 (e.g). There, you need to fix as many sigmas as possible, at least in my hands.
Tom, thanks for the density increment comment; that’s of course the bare-bones buoyancy, which is what the particle experiences. Keep it simple. Still: the vbar, as a composite parameter should be interpretable in “structural” terms (that might be accuracy-limited, I fear). I’ll ask our SAXS people about this. And yes, Heini Eisenberg as well, though insomnia is not my biggest concern right now.

Best to all,

Holger

From: Rufer, Arne [mailto:arne.rufer at roche.com]
Sent: torsdag, september 28, 2017 17:46
To: HGSR (Holger Martin Strauss)
Cc: rasmb at list.rasmb.org
Subject: Re: [RASMB] another vbar question

Hi Holger,

would preferential hydration play a role at the very high NaCl concentrations you are using? Regarding anecdotes on vbar, I can add that when we worked with acylated peptides the vbar measured by densitometry turned out to be much smaller than vbar calculated from the aa sequence + modification. We had congruent AUC and SEC-MALLS data showing that certain acylations led to oligomerisation, presumably due to an hydrophobic interaction that became stronger with higher salt concentration. At that time we argued the effect on vbar could be due to a solvent envelope for packed acyl-moieties that is smaller than that of the separated acyl-moieties. Makes sense? I am curious to read Mattia's comment on this, too.

Best,

Arne

On Thu, Sep 28, 2017 at 3:48 PM, HGSR (Holger Martin Strauss) <hgsr at novonordisk.com<mailto:hgsr at novonordisk.com>> wrote:
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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