I just tried that protocol, and had essentially all of my protein
crashed out. Any tips on optimizing the methylation reaction to
reduce precipitation? Perhaps reducing the formaldehyde or
dimethylaminoborane, shortening the incubation times, etc.?
Thanks,
Nat
On Wed, Apr 14, 2010 at 6:00 PM,
Maybe you could try a modification that does not require reduction?
Like sulfo-NHS-acetate (will turn primary amines into amides). There
are hundreds of amine-reactive protein modification reagents out
there. Most of them listed at piercenet.com.
Also, try doing the reaction at different
I was surprised to get a few messages asking for our protocol for
reductive methylation of proteins for crystallization. We employ almost
exactly the protocol published by Walter et al, Structure, 2006. This is
a Ways and Means article that made us realize how easy it was to do
this regularly
Hi all,
I'm currently trying to crystallise a two domain protein which
contains several structurally important disulfides. We have a high
resolution structure of one domain (~1.4 A resolution), which reveals
quite a few solvent-exposed lysines, some of which are involved in
Dear Oliver,
In our lab, reductive methylation using dimethylaminoborane is regularly
performed, and nearly everything we work on have native disulfides.
Among five or six reactions I've performed on molecules with disulfides,
I have not had a case where solubility or stability was affected.
