The Leishmania tarentolae exosome: purification and structural analysis by electron microscopy.
The eukaryotic exosome is a complex of at least 11 proteins that is required for various 3'-5' exoribonucleolytic RNA processing and degradation reactions. The minimal core consists of 6 RNase PH and 3 S1 domain subunits; various additional proteins may be associated. We describe here the purification of native exosome from Leishmania tarentolae. The yield is sufficient for structural studies of the native exosome. Electron microscopy and image reconstruction of negatively stained preparations revealed the expected six-membered ring structure at 35 A resolution. An additional density suggested that RRP6 and its partner EAP3 (equivalent to Rrp47) might be located at the top of the exosome and at the side of the hexameric ring. No exonuclease or polyadenylation activity was detected in the exosome preparations.
Cristodero
Marina
M
Zentrum für Molekulare Biologie der Universität Heidelberg (ZMBH), Im Neuenheimer Feld 282, D-69120 Heidelberg, Germany.
Böttcher
Bettina
B
Diepholz
Meikel
M
Scheffzek
Klaus
K
Clayton
Christine
C
D000818:Animals; D003001:Cloning, Molecular; D005095:Exoribonucleases / Q000737:chemistry* / Q000302:isolation & purification* / Q000378:metabolism; D007091:Image Processing, Computer-Assisted; D007891:Leishmania / Q000201:enzymology* / Q000648:ultrastructure; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D021122:Protein Subunits / Q000737:chemistry* / Q000302:isolation & purification* / Q000378:metabolism; D015800:Protozoan Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D016053:RNA, Protozoan / Q000378:metabolism
D021122:Protein Subunits; D015800:Protozoan Proteins; D012333:RNA, Messenger; D016053:RNA, Protozoan; D005095:Exoribonucleases