Proteomics

Dataset Information

0

Assembly defects of the human tRNA splicing endonuclease contribute to impaired pre-tRNA processing in pontocerebellar hypoplasia


ABSTRACT: To gain functional insights into human TSEN/CLP1, we designed an expression vector series based on the MultiBac system that allows combinatorial protein complex production in insect and mammalian cells by utilizing a CMV/p10 dual promoter. Using this system, we were able to assemble and purify functional heterotetrameric TSEN and a heteropentameric complex including the RNA kinase CLP1 from infected insect cells. Individual proteins of the complexes were identified using in-gel digestion and LC-MS/MS analysis. To gain atomistic insights into the molecular architecture of the human TSEN complex, we set out to characterize the TSEN15–34 heterodimer by X-ray crystallography. Despite extensive crystallization trials, full-length TSEN15–34 did not yield any crystals. To define a crystallizable core complex, we subjected the full-length complex to limited proteolysis with subsequent size exclusion chromatography. Primarily, we observed two comigrating polypeptide species. The two polypeptide species were identified using in-gel digestion followed by LC-MS/MS analysis. This led to the identification of the polypeptide species corresponding to residues 23 to 170 of TSEN15 and residues 208 to 310 of TSEN34 covering the predicted conserved nuclease domains.

INSTRUMENT(S): Q Exactive Plus

ORGANISM(S): Homo Sapiens (human)

SUBMITTER: Marie Alfes  

LAB HEAD: Carla Schmidt

PROVIDER: PXD019034 | Pride | 2021-06-17

REPOSITORIES: Pride

Dataset's files

Source:
Action DRS
2017_07_05_MB_3.raw Raw
2017_07_05_MB_4.raw Raw
2017_07_05_MB_5.raw Raw
2017_07_05_MB_6.raw Raw
2017_07_07_MB_31.raw Raw
Items per page:
1 - 5 of 25

Similar Datasets

2024-10-17 | PXD044729 | Pride
2024-10-17 | PXD044731 | Pride
2023-08-01 | GSE238196 | GEO
2018-11-07 | MODEL1811050001 | BioModels
2013-11-29 | E-GEOD-40197 | biostudies-arrayexpress
2024-10-17 | PXD044772 | Pride
2024-10-17 | PXD044742 | Pride
2024-10-17 | PXD044786 | Pride
2024-10-17 | PXD044788 | Pride
2024-10-17 | PXD044763 | Pride