Dear all,
Please, maybe you could give some suggestions to the problem below.
1) Images show smeared spots, but xds did a good job integrating them. The
cell is 229, 229, 72, trigonal, and we see alternating strong and weak rows
of spots in the images (spots near each other, but rows more separated, must
be by c*). They were scaled with xscale, P622 (no systematic abscences),
R_symm = 5.3 (15.1), I/sigI = 34 (14) and redundancy = 7.3 (6.8), resolution
2.8 A. Reciprocal space show strong spots at h, k, l=2n and weak spots at h,
k, l=2n+1 (I mean, l=2n intensities are practically all higher than l=2n+1
intensities, as expected from visual inspection of the images). Within
planes h, k, l=2n+1, the average intensity is clearly and "much" *higher at
high resolution than at low resolution*. Also, within planes h, k, l=2n, a
subjective observation is that average intensity apparently does not decay
much from low to high resolution. The data were trucated with truncate,
which calculated Wilson B factor to be 35 A**2.
2) Xtriage points a high (66 % of the origin) off-origin Patterson peak.
Also, ML estimate of overall B value of F,SIGF = 25.26 A**2.
3) I suspect to have a 2-fold NCS parallel to a (or b), halfway the c
parameter, which is "almost" crystallographic.
4) I submitted the data to the Balbes server which using
pseudo-translational symmetry suggested some solutions, one with a good
contrast to others, with a 222 tetramer, built from a structure with 40 %
identity and 58% positives, of a well conserved fold.
5) I cannot refine below 49 % with either refmac5, phenix.refine or CNS.
Maps are messy, except for rather few residues and short stretches near the
active site, almost impossible for rebuilding from thereby. Strange, to me,
is that all programs "freeze" all B-factors, taking them the program minimum
(CNS lowers to almost its minimum). Might this be due to by what I observed
in the reciprocal space as related in "1" ? If so, might my (intensity)
scaling procedure have messed the intensities due to their intrinsic
"property" to be stronger in alternating planes ? How to overcome this ?
6) I tried some different scaling strategies *in the refinement step*, no
success at all.
7) A Patterson of the solution from Balbes also shows an off-origin Patteron
at the same position of the native data, although a little lower.
8) Processed in P6, P312 and P321, all of course suggest twinning.
I would thank suggestions, point to similar cases, etc... In fact, currently
I wondered why refinement programs take B-factor to such low values
Many thanks,
Jorge
