Hi,
Ah yes, low resolution chain tracing.... I'm only in the initial
stages of map interpretation, but I anticipate I might have a similar
problem. I would love to hear from anyone who has ideas as well.
Thanks
Jeff
On Jun 15, 2007, at 10:03 AM, Alexei Datsuk wrote:
Hi,
I need some advice from experts. I have a low resolution electron
density map (3.7 angstroms) solved by SIRAS. The protein is a
tetramer (47 kDa/monomer) but I'm having trouble getting in
register. There is very good main chain density and clear
secondary structure (beta-sheets) so I likely have solved it
correctly. I can trace most of the main chain as fragments (loops
connecting beta strands are broken) but I can't tell if I'm tracing
it in the right direction or whether I have it backwards. There is
no real density for side chains at all (except for Calpha), not
even aromatics, so I have no clue which way is N or C-terminus.
There are a few disulfide bonds too so I'm real worried about
tracing incorrectly and never being able to fix it later. My
strategy right now is to build in as much poly-ala fragments (I can
get ~75% of trace as fragments) and then refine them. Then redo
phasing in SHARP with poly-ala model and SIRAS phases and then use
DM with NCS averaging to get improved map. Hopefully I can see
then side chain density. My question is how much bias would be
incorporated if some of the fragments are traced backwards? Would
I be able to sort things out if some fragments are traced
backwards? Has anyone traced chains as fragments not knowing which
direction is which and then be able to sort things out later?
Right now I am stuck- I have spent the last 1 months trying to get
things in register but no luck. I don't want to build haphazardly
but it doesn't look like I have much choice.
What have people done when tracing in low resolution maps?
Thanks
Alexei
