大肠杆菌K-12尿苷磷酸化酶的分离、在宏观重力场中的结晶及初步X射线研究。

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

作者信息

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

机构信息

Institute of Crystallography Academy of Sciences of the USSR, Moscow.

出版信息

Biochem Int. 1992 Mar;26(4):607-15.

PMID:1610370

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 of 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 P2(1)2(1)2(1) with the unit cell constants a = 90.4; b = 128.8; c = 136.8 A. There is one molecule per asymmetric unit, Vm = 2.4. These crystals diffract to at least 2.5-2.7 A resolution. The hexameric structure of UPH was directly demonstrated by electron microscopy study and image processing.

摘要

已从一个在多拷贝质粒上携带编码尿苷磷酸化酶（UPH）的udp基因的大肠杆菌K-12菌株中，将尿苷磷酸化酶（UPH）纯化至接近均一。UPH被纯化至电泳纯，比活性为230单位/毫克，回收率为80%，从3克细胞中获得了120毫克该酶。在制备型超速离心机中获得了适合X射线衍射分析的酶晶体。晶体中分子的堆积可由空间群P2(1)2(1)2(1)描述，晶胞常数a = 90.4；b = 128.8；c = 136.8 Å。每个不对称单位有一个分子，Vm = 2.4。这些晶体的衍射分辨率至少为2.5 - 2.7 Å。通过电子显微镜研究和图像处理直接证明了UPH的六聚体结构。