Dear Pu, sorry for the long(ish) answer ... handedness/enantiomorph, heavy atom sites and consistency (while using SHARP/autoSHARP) are just one of my favourites ;-)
On Fri, Aug 20, 2010 at 01:39:58PM +0100, Pu Gao wrote: > I rechecked the sharp logs, and found that the original SAD sites > were wrong, which had to be turned to its inverted ones for > calculating the final density (while the SIRAS sites are correct, > the final density was calculated directly from the original sites). If you run autoSHARP including the density modification step (!) it will try and detect which is the correct hand/enantiomorph. It is then important to either take the last SHARP run (inverted hand) or the last but one (original hand) when continuing to work with the current heavy atom model in SHARP (e.g. to then do MAD+native or spherical averaged heavy atom clusters etc). The correct set of heavy atom parameters should be clearly stated in the log file from autoSHARP. If you did run e.g. SHELXD by hand and then used those sites in SHARP you could obviously have the wrong hand/enantiomorph (see George's post). I would generally recommend using the heavy atom sites from SHELXD in an autoSHARP run: unless you do something more complex than SAD/MAD or SIR(AS)/MIR(AS). It will then skip it's own heavy-atom detection step (also using SHELXC/D) and use your supplied sites. Using sites from your own HA-detection attempt is a good idea for tricky cases (where you might have to run a lot of trials), cases where the number of sites is very uncertain and you need to run lots of slightly different trials ... or if you have very low resolution data/signal (5A and lower). In any case: make your life easy and plug them into autoSHARP up to the density modifcation step. Also, if you run several phasing scenaria (SAD, another one SIRAS, different derivatives etc) there is a very useful feature in SHARP that allows input of external (prior) phase information - e.g. in form of Hendrickson-Lattmann coefficients. This way you could use the SHARP phases from your SIRAS job (e.g. nat+Hg) for a SAD run (Se-MET peak - no sites known so far) and detect your Se sites in the log-likelihood gradient maps ... on the same origina (!) as the other sites. Having all sites on the same origina makes your life MUCH easier if you have to deal with lots of derivatives, datasets and slightly different cell dimensions. You could also use your phases from a poor or partial MR solution in the same way to detect your heavy atom sites. This feature (using prior phase information from partial models, MR or other derivatives) has been in SHARP since the very early days and has been useful to us in a lot of cases. See e.g. the 2010 paper by Pietro Roversi (http://scripts.iucr.org/cgi-bin/paper?ba5141) or some earlier references: Kauppi B, Lee K, Carredano E, Parales RE, Gibson DT, Eklund H, Ramaswamy S. (1998). Structure of an aromatic-ring-hydroxylating dioxygenase-naphthalene 1,2-dioxygenase. Structure 6, 571-86. and the manual: http://www.globalphasing.com/sharp/manual/chapter2.html#external http://www.globalphasing.com/sharp/manual/chapter4.html#ExternalPhaseInformation Cheers Clemens -- *************************************************************** * Clemens Vonrhein, Ph.D. vonrhein AT GlobalPhasing DOT com * * Global Phasing Ltd. * Sheraton House, Castle Park * Cambridge CB3 0AX, UK *-------------------------------------------------------------- * BUSTER Development Group (http://www.globalphasing.com) ***************************************************************
