Isolation, crystallization in the macrogravitation field, preliminary x-ray investigation of Uridine phosphorylase from Escherichia coli K-12.

A. M. Mikhailov, E. A. Smirnova, V. L. Tsuprun, I. V. Tagunova, B. K. Vainshtein, E. V. Linkova, A. A. Komissarov, Z. Z. Siprashvili, A. S. Mironov

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Abstract

Uridine phosphorylase (UPH) from Escherichia coli K-12 has been purified to near homogeneity from a strain harbouring the udp gene, encoding UPH, on a multicopy plasmid. UPH was purified to electrophoretic homogeneity with the specific activity 230 units/mg with a recovery of 80%, yielding 120 mg of enzyme from 3g cells. Crystals or enzyme suitable for X-ray diffraction analysis were obtained in a preparative ultracentrifuge. The packing of the molecules in the crystals may be described by the space group P212121 with the unit cell constants a=90.4; b=128.8; c=136.8 A. There is one molecule per asymmetric unit, V(m)=2.4. These crystals diffract to at least 2.5 - 2.7 Å resolution. The hexameric structure or UPH was directly demonstrated by electron microscopy study and image processing.

Original languageEnglish (US)
Pages (from-to)607-615
Number of pages9
JournalBiochemistry International
Volume26
Issue number4
StatePublished - Jan 1 1992

Keywords

  • Cryatallization
  • Uridine phosphorylase
  • X-ray analysis

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