2M5X
Novel method of protein purification for structural research. Example of ultra high resolution structure of SPI-2 inhibitor by X-ray and NMR spectroscopy.
Summary for 2M5X
| Entry DOI | 10.2210/pdb2m5x/pdb |
| Related | 4hgu |
| NMR Information | BMRB: 19085 |
| Descriptor | Silk protease inhibitor 2 (1 entity in total) |
| Functional Keywords | serine protease inhibitor, kazal family, spi-2, hydrolase inhibitor |
| Biological source | Galleria mellonella (Greater wax moth) |
| Total number of polymer chains | 1 |
| Total formula weight | 4317.74 |
| Authors | Lenarcic Zivkovic, M.,Dvornyk, A.,Kludkiewicz, B.,Kopera, E.,Zagorski-Ostoja, W.,Grzelak, K.,Zhukov, I.,Bal, W. (deposition date: 2013-03-12, release date: 2014-03-12, Last modification date: 2024-10-30) |
| Primary citation | Kopera, E.,Bal, W.,Lenarcic Zivkovic, M.,Dvornyk, A.,Kludkiewicz, B.,Grzelak, K.,Zhukov, I.,Zagorski-Ostoja, W.,Jaskolski, M.,Krzywda, S. Atomic resolution structure of a protein prepared by non-enzymatic His-tag removal. Crystallographic and NMR study of GmSPI-2 inhibitor. Plos One, 9:e106936-e106936, 2014 Cited by PubMed Abstract: Purification of suitable quantity of homogenous protein is very often the bottleneck in protein structural studies. Overexpression of a desired gene and attachment of enzymatically cleavable affinity tags to the protein of interest made a breakthrough in this field. Here we describe the structure of Galleria mellonella silk proteinase inhibitor 2 (GmSPI-2) determined both by X-ray diffraction and NMR spectroscopy methods. GmSPI-2 was purified using a new method consisting in non-enzymatic His-tag removal based on a highly specific peptide bond cleavage reaction assisted by Ni(II) ions. The X-ray crystal structure of GmSPI-2 was refined against diffraction data extending to 0.98 Å resolution measured at 100 K using synchrotron radiation. Anisotropic refinement with the removal of stereochemical restraints for the well-ordered parts of the structure converged with R factor of 10.57% and Rfree of 12.91%. The 3D structure of GmSPI-2 protein in solution was solved on the basis of 503 distance constraints, 10 hydrogen bonds and 26 torsion angle restraints. It exhibits good geometry and side-chain packing parameters. The models of the protein structure obtained by X-ray diffraction and NMR spectroscopy are very similar to each other and reveal the same β2αβ fold characteristic for Kazal-family serine proteinase inhibitors. PubMed: 25233114DOI: 10.1371/journal.pone.0106936 PDB entries with the same primary citation |
| Experimental method | SOLUTION NMR |
Structure validation
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