Just to add to what has been "said" (written) before:
coiled coiled or simply helices can be very problematic for M.R.. Human
sacrifice has never given any positive result (as reported in the
literature as far as I know), but "heavy atom sacrifice" could be
attempted ("heavy atom" includes in my view atoms that are used for SAD,
MAD - such as S, Se... - in addition to Au, Pt...) in parallel to your
M.R. searches which may never provide you with an acceptable solution.
Fred.
Napoleão Valadares wrote:
Hi there!
I got crystals from some synthetic peptides I bought, they are 30
residues long and are supposed to form a coiled coil. I collected
various data sets (home source, Brookhaven and Diamond), including
some at the resolution of 1.65 A, for which the space group appears to
be C222 or C2221. The unit cell is small, 22.67, 88.06, 26.13, and the
Matheus Coefficient indicates that's there's only one helix in the
asymmetric unit and a 25% solvent content.
I have tried A LOT of Molecular Replacement using Phaser and
Phenix AutoMR. I'm using a 80% identity coiled coil helix as search
model. The programs give me solutions with "reasonable" maps, but it
is never possible to refine to achieve Rvalues below 0.40.
Additionally, maps from different solutions look reasonable, so I'm
thinking these are all bias.
I have 5 other synthetic 30 residues peptides (that crystallize in
different space groups and diffract to lower resolutions), including a
SelenoMethionine (SM) derivative (but it does not have enough
anomalous signal, ASU is too big, it is possible that the SM are
disordered). I'm stuck on this since March.
Regarding the search model, I already tried trimming some or all
side chains and removing 2, 3 or 5 residues on each/both sides. I also
tried other search models. Maybe some "magic" combination of
parameters on Phaser or other programs can help me.
What is your advice regarding how to proceed with MR? Is there
some program, procedure, parameter, pray or human sacrifice that could
help me?
Thank you.
Regards,
Napo
