Dear all, Sorry for the non-CCP4 question.
We recently crystallized a protein expressed from pGEX4T-1 vector. The GST tag is cleaved by thrombin before crystallization. The cleavage site of thrombin is Leu-Val-Pro-Arg || Gly-Ser, thus Gly and Ser should remain at the N-terminus of the protein. Strangely, from the 2.5A map, we observed some unexplainable electron density before the most N-terminal glycine residue (http://www.dddc.ac.cn/group/htc/NT_map.jpg). Has anyone encountered similar situation with thrombin cleavage? Could it be some kind of modification or additional residues resulted from unspecific cleavage of thrombin? The crystallization liquor contains MES, PEG6000, Tris, DTT, NaCl and EDTA. Any suggestion is highly appreciated! Thanks in advance! -------------- Tiancen Hu Shanghai Institute of Materia Medica Chinese Academy of Sciences Rm. 2107, #555, ZuChongzhi Rd. Shanghai 201203 P.R. China Phone: +86-21-50806600 ext. 2107
