Crystallization of the Bacillus subtilis SPP1 bacteriophage helicase loader protein G39P
The essential helicase loader protein G39P encoded by Bacillus subtilis SPP1 phage has been overproduced in Escherichia coli and purified. The wild‐type protein has been crystallized by the hanging‐drop vapour‐diffusion method in a primitive hexagonal space group, probably P6122/P6522, but the cryst...
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Published in | Acta crystallographica. Section D, Biological crystallography. Vol. 59; no. 6; pp. 1090 - 1092 |
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Main Authors | , , , , , |
Format | Journal Article |
Language | English |
Published |
5 Abbey Square, Chester, Cheshire CH1 2HU, England
Munksgaard International Publishers
01.06.2003
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Subjects | |
Online Access | Get full text |
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Summary: | The essential helicase loader protein G39P encoded by Bacillus subtilis SPP1 phage has been overproduced in Escherichia coli and purified. The wild‐type protein has been crystallized by the hanging‐drop vapour‐diffusion method in a primitive hexagonal space group, probably P6122/P6522, but the crystals diffract to only 3.4 Å and are poorly reproducible. Mass‐spectrometric analysis has revealed marked proteolytic cleavage from the C‐terminus and the presence of a major species corresponding to deletion of the 14 C‐terminal residues. Thus, a new variant of the protein (G39P112) has been engineered that corresponds to a 14‐residue C‐terminal truncation. The G39P112 variant has also been crystallized but now in a primitive orthorhombic form, probably P21212 or P212121, with unit‐cell parameters a = 85.6, b = 89.7, c = 47.6 Å, with diffraction to 2.4 Å on a synchrotron source and with greatly improved reproducibility. Calculation of VM values for this G39P112 variant suggests the presence of three monomers in the asymmetric unit, corresponding to a solvent content of about 47%. A selenomethionine‐incorporated form of the protein has been produced and a full three‐wavelength MAD data collection undertaken. |
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Bibliography: | ArticleID:AYDCY0087 istex:76598B3A2809860AA23FAAEC4E0AA5A2E4327631 ark:/67375/WNG-DBJKXRRP-P ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23 |
ISSN: | 1399-0047 0907-4449 1399-0047 |
DOI: | 10.1107/S0907444903007236 |