Production and crystallization of a selenomethionyl variant of UmuD', an Escherichia coli SOS response protein.
Crystals of both native and mutant Escherichia coli UmuD' protein were obtained using the hanging drop method. Soaking the native crystals in solutions of heavy metal ions failed to produce good isomorphous derivatives, and selenomethionine substituted wild-type protein did not crystallize under conditions that gave native crystals. Site-directed mutagenesis was used to change the penultimate residue, a methionine amino acid, to either a valine or a threonine amino acid. Crystals were subsequently obtained from these mutant proteins with and without selenomethionine incorporation. Crystals of the native, the mutant, and the selenomethionine incorporated protein were all similar, crystallizing in the P4(1)2(1)2 space group.
Peat
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TS
Department of Biochemistry and Molecular Biophysics, Columbia University, New York, New York 10032, USA.
Frank
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EG
Woodgate
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Hendrickson
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WA
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D018360:Crystallography, X-Ray; D004259:DNA-Directed DNA Polymerase; D029968:Escherichia coli Proteins; D008969:Molecular Sequence Data; D016296:Mutagenesis; D016297:Mutagenesis, Site-Directed; D016133:Polymerase Chain Reaction; D017386:Sequence Homology, Amino Acid
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D004259:DNA-Directed DNA Polymerase; C055218:UmuD protein, E coli