7NJH
HEX1 (in cellulo) grown inside HARE serial crystallography chip
Summary for 7NJH
| Entry DOI | 10.2210/pdb7njh/pdb |
| Descriptor | Woronin body major protein (2 entities in total) |
| Functional Keywords | serial crystallography, hex1, in-cellulo, silicon chip, structural protein |
| Biological source | Neurospora crassa (strain ATCC 24698 / 74-OR23-1A / CBS 708.71 / DSM 1257 / FGSC 987) |
| Total number of polymer chains | 1 |
| Total formula weight | 19150.66 |
| Authors | Norton-Baker, B.,Mehrabi, P.,Boger, J.,Schonherr, R.,von Stetten, D.,Schikora, H.,Martin, R.W.,Miller, R.J.D.,Redecke, L.,Schulz, E.C. (deposition date: 2021-02-16, release date: 2021-06-16, Last modification date: 2024-01-31) |
| Primary citation | Norton-Baker, B.,Mehrabi, P.,Boger, J.,Schonherr, R.,von Stetten, D.,Schikora, H.,Kwok, A.O.,Martin, R.W.,Miller, R.J.D.,Redecke, L.,Schulz, E.C. A simple vapor-diffusion method enables protein crystallization inside the HARE serial crystallography chip. Acta Crystallogr D Struct Biol, 77:820-834, 2021 Cited by PubMed Abstract: Fixed-target serial crystallography has become an important method for the study of protein structure and dynamics at synchrotrons and X-ray free-electron lasers. However, sample homogeneity, consumption and the physical stress on samples remain major challenges for these high-throughput experiments, which depend on high-quality protein microcrystals. The batch crystallization procedures that are typically applied require time- and sample-intensive screening and optimization. Here, a simple protein crystallization method inside the features of the HARE serial crystallography chips is reported that circumvents batch crystallization and allows the direct transfer of canonical vapor-diffusion conditions to in-chip crystallization. Based on conventional hanging-drop vapor-diffusion experiments, the crystallization solution is distributed into the wells of the HARE chip and equilibrated against a reservoir with mother liquor. Using this simple method, high-quality microcrystals were generated with sufficient density for the structure determination of four different proteins. A new protein variant was crystallized using the protein concentrations encountered during canonical crystallization experiments, enabling structure determination from ∼55 µg of protein. Additionally, structure determination from intracellular crystals grown in insect cells cultured directly in the features of the HARE chips is demonstrated. In cellulo crystallization represents a comparatively unexplored space in crystallization, especially for proteins that are resistant to crystallization using conventional techniques, and eliminates any need for laborious protein purification. This in-chip technique avoids harvesting the sensitive crystals or any further physical handling of the crystal-containing cells. These proof-of-principle experiments indicate the potential of this method to become a simple alternative to batch crystallization approaches and also as a convenient extension to canonical crystallization screens. PubMed: 34076595DOI: 10.1107/S2059798321003855 PDB entries with the same primary citation |
| Experimental method | X-RAY DIFFRACTION (2.5 Å) |
Structure validation
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