I'm dealing with peptides which are on one hand polar (much lysine content) and on the other they have a fatty acid acylation. This causes problems regarding the attachment of these peptides on various surfaces. Has anyone had experiences with such peptides and what could help to avoid these unwanted sticking effects?
I already tried glassware which does not work at all. I tried low affinity binding materials, which is useful, however there is still a lot of loss. Are there any suitable buffer/solvent mixtures which can further prevent the adsorption?
Hi Tom,
if OK with your assay, you may want to increase the amount of carrier protein (BSA usually) to up to 5%. We have successfully been using 2% BSA with very hydrophobic peptides to reduce unspecific binding in functional GPCR assays.
Best regards,
Carsten
I've found that methanol is a good solvent for such materials. Addition of detergents like SDS or DPC (dodecylphosphatidyl choline) also helps.
What quantity are you dealing with? I'm used to mg quantities for NMR.
Hi
In the protocols for MIC testing for example, we are always working with amphiphilic peptide or peptoids. Before we are working with them in biological assays on polypropylene or polystyrene plates we are dissolving stock concentrations in solvent containing o.2% BSA and 0.01% acetic acid. We are also using polypropelene plates to avoid sticking of the peptides to the plate.
Hope this helps
Hi Tom!
I have for many years used a combination of 2% BSA and 1% Triton-X100 in phosphate buffers for "sticky" peptides.
Best regards
Elvar
I agree, that the answer depends on what you intend to do with it. But try 5% DMSO or less.
First, many thanks to all of you for your great answers.
I want to purify these amphiphilic peptides from biologic fluids and want to analyze them by LC-MS. So 5%DMSO or less, should be okay. However, unfortunately I cannot use MeOH:Chl (50:50), because my compound would not interfere with the LC-column material anymore.
Anyway, I will try your suggestions in an adjusted version.
Thanks again.
Tom
Dear Tom,
You can then use Acetonitrile:tert-butanol: Water (1:1:1). You can also use it for LC-MS. I am sure this will work for you. Could you please tell in detail, which peptide and which lipid?
Harshal
I've had similar issues... To avoid it, I make a 100x concentrated stock of my peptides in 80% DMSO and dilute to 1x in a solution containing 0.2% BSA.
I have worked with both amphiphilic peptides and purified proteins,Try using
silica-coated glassware.
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