Generally fab geometry presents problems for mr due to variable interdomain angle. Assuming your sg is correct and this is not a twin, i would try searching at 4 or even 5 a resolution. I would not reject the chance that you have high solvent content, so you might have fewer molecules in au than expected from vm estimates. We recently solved a whole bunch of structures with large unit cell dimensions and high solvent content in the 75% or even 82% range. All had issues with resolution, freezing, and twinning - similar to shat you describe. In a few cases we screened over 35 crystals (of the same protein variant, same condition, just repeats) to find one that diffracted beyond 4a. Don't give up! May the pork be with you.

Artem

On Dec 9, 2014 12:32 PM, "Mo Wong" <[log in to unmask]> wrote:

Hi all,

I’m stuck on a rather complex molecular replacement problem. The crystals are of an antigen-Fab complex totaling ~67 kDa (waiting to confirm using PAGE gel). They diffract to ~3.5A at the synchrotron and process in C2 with dimensions 220x130x230 and beta at 103 so it looks like there are round 9to12-ish copies in the ASU. The overall Rmerge is high at ~25% with I/sig cutoff ~2 and redundancy of 5; however, at 4.5A this drops to ~15%. Furthermore, processing in P1 gives similar Rmerge values too.

Self-Patterson doesn’t suggest translational symmetry, but the self-rotation function (SRF) suggests high NCS (see below/attached).

I’m hoping the SRF might be helpful in trying to confirm/dismiss C2 is the likely space group, and perhaps suggest a logical assembly with the ASU (I see strong 2-fold and 3-fold NCS operators suggesting to me dimeric trimers or vice versa - however, I’ve never had to really analyze SRFs in the absence of a mol. rep. solution so my interpretation could be wrong).

Anyway, any help to bringing a molecular replacement solution closer to reality would be appreciated.

Thanks!