My name is Greg Costakes and I am a graduate student for Cynthia Stauffacher at Purdue University. I am attempting to solve the structure of an 18kDa protein domain using molecular replacement to obtain proper phasing (44% identical / 78% homologous to an existing structure). Crystals diffracted to roughly 2 angstroms and are in the space group P212121. There are 10,200 unique reflections with 5.7 fold redundancy. I chose to make an R(free) set containing 10% of the reflections so that I will have just over 1000 in the set.
After failing to obtain a valid solution from Phaser and Molrep, I turned to BALBES which was able to successfully perform the molecular replacement and give me final R/R(free) values of 0.3/0.34. However, I noticed the MTZ file that BALBES gave me contained an R(free) set of 5% containing only 500 reflections. I was wondering why BALBES changed the percentage of reflections pegged for the R(free) set. Is there a way to prevent BALBES from changing your R(free) fraction size? Also, is 500 reflections enough for an R(free) set? I had previously been told that R(free) should contain between 1000-1500 reflections. Please let me know. Thank you. ------------------------------------------------------------------------------- Greg Costakes PhD Candidate Department of Structural Biology Purdue University Hockmeyer Hall, Room 320 240 S. Martin Jischke Drive, West Lafayette, IN 47907 --------------------------------------------------------------------------------
