Object's details: Atomic resolution structure of a protein prepared by non-enzymatic His-tag removal. Crystallographic and NMR study of GmSPI-2 inhibitor.

Provider:Repozytorium Instytutu Biochemii i Biofizyki Polskiej Akademii Nauk

Description

Title:
Atomic resolution structure of a protein prepared by non-enzymatic His-tag removal. Crystallographic and NMR study of GmSPI-2 inhibitor.
Creator:
Kopera, Edyta;
Bal, Wojciech;
Lenarčič Živković, Martina;
Dvornyk, Angela;
Kludkiewicz, Barbara;
Grzelak, Krystyna;
Zhukov, Igor;
Zagórski-Ostoja, Włodzimierz;
Jaskolski, Mariusz;
Krzywda, Szymon
Description:
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.
Object availability:
free access
Date:
2014
Type:
Article;
PeerReviewed
Language:
English
Format:
application/pdf
Relation: http://eprints.ibb.waw.pl/797
Subject:
Q Science (General);
QD Chemistry;
QH301 Biology
Identifier: http://eprints.ibb.waw.pl/797/1/journal.pone.0106936.pdf ;
Kopera, Edyta and Bal, Wojciech and Lenarčič Živković, Martina and Dvornyk, Angela and Kludkiewicz, Barbara and Grzelak, Krystyna and Zhukov, Igor and Zagórski-Ostoja, Włodzimierz and Jaskolski, Mariusz and Krzywda, Szymon (2014) Atomic resolution structure of a protein prepared by non-enzymatic His-tag removal. Crystallographic and NMR study of GmSPI-2 inhibitor. PloS one, 9 (9). e106936. ISSN 1932-6203
Data provider:
Repozytorium Instytutu Biochemii i Biofizyki Polskiej Akademii Nauk

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Object's details: Atomic resolution structure of a protein prepared by non-enzymatic His-tag removal. Crystallographic and NMR study of GmSPI-2 inhibitor.

Description

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