Wouldn't a HSQC of 15N-labeled protein be a relatively easy yes/no experiment? Maybe it would not be incredibly definitive?
Jacob On Mon, Dec 6, 2010 at 11:10 AM, Mischa Machius <[email protected]> wrote: > Daniel, > > You'll probably have to monitor pH changes through size changes of your > protein, provided the structural changes will indeed cause size changes. > > You said "easy", so that probably rules out Small-Angle X-Ray Scattering > (SAXS), but that would be the highest-resolution method. You can try static > and dynamic light scattering, analytical ultracentrifugation and > fluorescence anisotropy. If you are really lucky, size exclusion > chromatography might work too. > > And then there are the "difficult" ways... > > MM > > > > > On Dec 6, 2010, at 11:59 AM, Daniel Jin wrote: > > Dear CCP4 colleagues, > > > > We have a protein that is composed of two domains connected by a short > peptide linker. We have some indirect evidence showing that the two domains > may somehow move against each other when exposed to different pH. It is > unlikely to have any obvious secondary structure change since each domain > behaves like a rigid body. I am wondering whether there is any “easy” way, > biochemically or biophysically, to monitor the conformational changes in > solution. Many thanks. > > > > As far as I know most of the pH sensing stories are linked to histidine > residue. Can you point me to any references that show a different pH sensing > mechanism (other than His)? Thanks. > > > > Best, > > Daniel > > > ----------------------------------------------------------------------- > Mischa Machius, PhD > Director, Center for Structural Biology > Assoc. Professor, Dept. of Pharmacology > Member, Lineberger Comprehensive Cancer Center > University of North Carolina > 4079 Genetic Medicine > CB#7365 > 120 Mason Farm Road > Chapel Hill, NC 27599-7365, U.S.A. > tel: +1-919-843-4485 > fax: +1-919-966-5640 > email: [email protected] <[email protected]> > > -- ******************************************* Jacob Pearson Keller Northwestern University Medical Scientist Training Program cel: 773.608.9185 email: [email protected] *******************************************
