Purification of U3 snoRNA Binding Homologs from Two Archaeal Organixms
Loading...
Authors
Long, Robert N.
Issue Date
2002
Type
Thesis
Language
en_US
Keywords
Alternative Title
Abstract
Small nuclear ribonucleoparticles (snRNPs) are a class of protein-RNA complexes found in the nucleus of eukaryotic cells. snRNPs are essential particles involved in the post-transcriptional processing of RNA. A subclass of snRNPs, the small nucleolar ribonucleoparticles (snoRNPs), are localized to the nucleolus and are involved in processing of the rRNA precursor transcript (pre-rRNA) and ribosome biogenesis. The U3 snoRNP is an essential snoRNP involved in the cleavage and methylation of pre-rRNA. These events are critical for the proper assembly of the ribosome. Four proteins, Nop1p, Nop56p, Nop58p and Snu13p have been shown to bind to the
U3 snoRNA, and form the U3 core complex. Because large protein complexes isolated from archaeal sources seem to be more amenable to crystallization than those from eukaryotic sources, it may be useful to study the structure and function of the archaeal U3 snoRNP. While no U3 complex has been observed in Archaea, proteins homologous to many U3 proteins do exist in Archaea. We have amplified genes encoding the U3 proteins Nop1, Nop56/58 from two archaeal genomes, S. solfataricus and M.jannaschii, as well as a Snu13 homolog from Mjannaschii.
These genes have been cloned into prokaryotic expression vectors and expressed in E. coli cells.
Recombinant Nop1p, Nop56/58p and Snu13p proteins will be purified by heat treatment and column chromatography. They will then be assayed for complex formation and for assembly upon the U3 snoRNA. These complexes will be purified and used in crystal trials for the determination of the crystal structure. This structure will provide some insights into the role of the U3 complex in pre-rRNA processing and will provide the basis for a structural and functional comparison of rRNA processing between Archaea and Eukarya.
Description
vi, 34 p.
Citation
Publisher
Kalamazoo College
License
U.S. copyright laws protect this material. Commercial use or distribution of this material is not permitted without prior written permission of the copyright holder.