Characterization of four orthologs of stringent starvation protein A.
Orthologous proteins can be beneficial for X-ray crystallographic studies when a protein from an organism of choice fails to crystallize or the crystals are not suitable for structure determination. Their amino-acid sequences should be similar enough that they will share the same fold, but different enough so that they may crystallize under alternative conditions and diffract to higher resolution. This multi-species approach was employed to obtain diffraction-quality crystals of the RNA polymerase (RNAP) associated stringent starvation protein A (SspA). Although Escherichia coli SspA could be crystallized, the crystals failed to diffract well enough for structure determination. Therefore, SspA proteins from Yersinia pestis, Vibrio cholerae and Pseudomonas aeruginosa were cloned, expressed, purified and subjected to crystallization trials. The V. cholerae SspA protein failed to crystallize under any conditions tested and the P. aeruginosa SspA protein did not form crystals suitable for data collection. On the other hand, Y. pestis SspA crystallized readily and the crystals diffracted to 2.0 A.
Andrykovitch
Michelle
M
Macromolecular Crystallography Laboratory, National Cancer Institute, National Institutes of Health, Frederick, MD 21702, USA.
Routzahn
Karen M
KM
Li
Mi
M
Gu
Yijun
Y
Waugh
David S
DS
Ji
Xinhua
X
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics; D003001:Cloning, Molecular; D003460:Crystallization; D004926:Escherichia coli / Q000737:chemistry / Q000235:genetics; D029968:Escherichia coli Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics; D008969:Molecular Sequence Data; D011550:Pseudomonas aeruginosa / Q000737:chemistry / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D014734:Vibrio cholerae / Q000737:chemistry / Q000235:genetics; D014961:X-Ray Diffraction; D015010:Yersinia pestis / Q000737:chemistry / Q000235:genetics
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D011994:Recombinant Proteins; C072071:SspA protein, E coli