XendoU is the first endoribonuclease described in higher eukaryotes as being involved in the endonucleolytic processing of intron-encoded small nucleolar RNAs. It is conserved among eukaryotes and its viral homologue is essential in SARS replication and transcription. The large-scale purification and crystallization of recombinant XendoU are reported. The tendency of the recombinant enzyme to aggregate could be reversed upon the addition of chelating agents (EDTA, imidazole): aggregation is a potential drawback when purifying and crystallizing His-tagged proteins, which are widely used, especially in highthroughput structural studies. Purified monodisperse XendoU crystallized in two different space groups: trigonal P3121, diffracting to low resolution, and monoclinic C2, diffracting to higher resolution.
Large-scale purification and crystallization of the endoribonuclease XendoU: troubleshooting with His-tagged proteins
Arceci M;Bozzoni I;Laneve P;Caffarelli E
2006
Abstract
XendoU is the first endoribonuclease described in higher eukaryotes as being involved in the endonucleolytic processing of intron-encoded small nucleolar RNAs. It is conserved among eukaryotes and its viral homologue is essential in SARS replication and transcription. The large-scale purification and crystallization of recombinant XendoU are reported. The tendency of the recombinant enzyme to aggregate could be reversed upon the addition of chelating agents (EDTA, imidazole): aggregation is a potential drawback when purifying and crystallizing His-tagged proteins, which are widely used, especially in highthroughput structural studies. Purified monodisperse XendoU crystallized in two different space groups: trigonal P3121, diffracting to low resolution, and monoclinic C2, diffracting to higher resolution.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
