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Title
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Expression, purification, crystallization and preliminary diffraction data characterization of Escherichia coli ribonuclease II (RNase II).
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Authors
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C.E.McVey,
M.Amblar,
A.Barbas,
F.Cairrão,
R.Coelho,
C.Romão,
C.M.Arraiano,
M.A.Carrondo,
C.Frazão.
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Ref.
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Acta Crystallograph Sect F Struct Biol Cryst Commun, 2006,
62,
684-687.
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PubMed id
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Abstract
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RNA degradation is important in the post-transcriptional control of gene
expression. The processing, degradation and quality control of RNA is performed
by many different classes of ribonucleases. Ribonuclease II (RNase II) is a
643-amino-acid enzyme that degrades single-stranded RNA from its 3'-end,
releasing ribonucleoside 5'-monophosphates. RNase II was expressed both as the
wild type and as a D209N mutant form. The latter was also produced as an SeMet
derivative. The various protein forms were crystallized using the
vapour-diffusion method. Wild-type RNase II was crystallized in two crystal
forms, both of which belonged to space group P2(1). X-ray diffraction data were
collected to 2.44 and 2.75 angstroms resolution, with unit-cell parameters a =
56.8, b = 125.7, c = 66.2 angstroms, beta = 111.9 degrees and a = 119.6, b =
57.2, c = 121.2 angstroms, beta = 99.7 degrees, respectively. The RNase II D209N
mutant gave crystals that belonged to space group P6(5), with unit-cell
parameters a = b = 86.3, c = 279.2 angstroms, and diffracted to 2.74 angstroms.
Diffraction data from the mutant and its SeMet derivative enabled the
determination of a partial Se-atom substructure by SIRAS.
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