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Dear all, we are trying to crystallize a protein (40 kD) that was cloned de novo as construct of five identical repeat structures. The protein contains no LYS, ARG, GLN, GLU, CYS, MET but a large number of negative charges (pI 3.7). The protein appears properly folded from CD, size exclusion. Crystallization hasn't worked out at 10 - 50 mg/ml. We also tried 10 - 50 mg/ml in presence of Ca and Zn, with and without Histag, we also added small diamines. Nothing worked so far. Recloning and mutational changes are not easy due the repetitive gene structure, however they are possible as the last anchor (including gene synthesis). 1. As we are very much interested in the protein structure I wonder whether people from other labs have any experience with highly charged proteins? 2. Are there any proteins with a significant surplus of positive charges crystallized that might be co-crystallized? 3. Did people use any chemicals to modify ASPs or GLUs side chains? Best wishes and thanks for your help! Kornelius ---------------------------------------------- Kornelius Zeth Max Planck Institute for Developmental Biology Dept. Protein Evolution Spemannstr. 35 72076 Tuebingen, Germany [EMAIL PROTECTED] Tel -49 7071 601 323 Fax -49 7071 601 349
