Hi, A few things you might try (or that you may have tried, but didn't mention):
1. Run the search models through chainsaw with their sequence alignments before searching (or just use poly-alanine versions of the search models), and reset the B-factor of the search models to the B-factor of the dataset (or your favorite number, the important part is that it's constant). 2. You mentioned that you get a solution for one domain. Is this a single Rotational/Translational solution set for 6 copies of this one search model, or something else? Assuming it is, you could try with fewer copies of your search model. If there are in fact 6 copies, and you search for 4, there should be density for all 6 in the maps phased from from the 4-model solution (assuming the MR solution is correct); but if it's the other way around there may be problems finding a solution. You may also want to try a range of combinations for searching (1 domain A + 1 domain B, 2 domain A + 1 domain B, 1 domain A + 2 domain B, etc). 3. It might be better not to bother with density modification for a 3.3 Angstrom model-phased dataset, especially if you have the possibility of using NCS. Good luck, Pete -----Original Message----- From: CCP4 bulletin board on behalf of Anjali Mehta Sent: Thu 3/6/2008 7:47 PM To: [email protected] Subject: [ccp4bb] Molecular replacement of a multidomain protein Dear All, I am working with a Bifunctional protein of molecular weight ~60 kDa. I have a 3.3 angstrom native dataset. The matthews number show there are 6 molecules in the asymmetric unit. The structures of the individual domains are already known from prokaryotes. The sequence identity with the known structures are about 30%. I have tried molecular replacement using the two parts as models respectively with CNS, MOLREP, PHASER etc. However I always get the solution for one domain. I have also tried to fix that domain and find the other one. But none of the programs can find a solution. I am trying to model build the correct sequence of one domain using a density modified (using CNS), NCS averaged (using RAVE) map but the map does not look very good. The side chains are not clear. That might be due to the fact that I am only having a partial model. Any suggestion will be appreciated. Thanks. Ms. Anjali Mehta
