Hi -- thank you for all your help. The majority opinion seems to be a
metal for the sphere (Ni from the Ni-affinity column, which (Joe
Patel, correct) was used during purification, but Zn and Fe were also
mentioned), and either water molecules, bis-tris or some other small
molecule forming the crescent. Just looking at the density, the
occupancy would seem to be quite high, so I'm surprised that a Ni ion
(or a contaminating metal ion) could have gone through the
purification and still remained at high enough concentration to be
clearly visible in the crystals. However, I'll still try this but
first some points of clarification and questions which you can either
email me seperately or post to the the group.
a. it was collected at beamline 19-BM at Argonne, so radiation damage
is an issue. Thierry Fishmann -- for the gln residue, there was
difference density for the gamma carbon after the first conformation
was modeled in, thus the addition of the second conformation, which I
agree is suspect. What does the radiation damage do chemically and
would that make the gamma carbon more mobile?
b. Jeffrey D Brodin -- how did you model in the bis-tris? Looking at
the bis-tris molecule from Hic-up, was the N at the centre of the
crescent and the O6 and O8 at the edges?
c. JR Helliwell -- there are 4 molecules in the AU, but two H138's
are pointing into the solvent. Thus the molar ratio of protein
molecules to "thing 1" is 4:1. Also looking at the images, I see no
ice rings -- the images look pretty good. Can you tell me more about
the series termination effects?
Again thank you for your help and I'll let the group know how it worked out.
Alex
--
Dr. Alex Singer
C.H. Best Institute
112 College St. Room 70
University of Toronto
Toronto, Canada, M5G 1L6
416-978-4033
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