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Title: Mapping the Arabidopsis organelle proteome
Author: Dunkley, Thomas Peter John
Awarding Body: University of Cambridge
Current Institution: University of Cambridge
Date of Award: 2006
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The work presented in this thesis involves the development of a proteomic method for protein localization that is not dependent on the preparation of pure organelles. This method has subsequently been named localization of organelle proteins by isotope tagging (LOPIT). Organelles are first partially separated using centrifugation through density gradients. Distributions of proteins within such gradients are then assessed by measuring the relative abundance of proteins between fractions along the length of the gradients. This is achieved using isotope-coded tags for protein quantitation by mass spectrometry. The subcellular localizations of proteins can then be determined by comparing their distributions to those of previously localized proteins, since proteins that belong to the same organelle will co-fractionate in the density gradients. Application of the LOPIT technique to the study of the Arabidopsis endomembrane system has resulted in the simultaneous assignment of 527 proteins to the endoplasmic reticulum, Golgi apparatus, vacuole, plasma membrane or to the mitochondria and plastids. These results demonstrate that proteomic analysis of the major endomembrane components can be performed in parallel, enabling protein steady state distribution between these organelles to be determined. Consequently, for the first time by proteomics, genuine residents of the endoplasmic reticulum, Golgi apparatus, vacuole and plasma membrane have been distinguished from contaminants and proteins that are in transit through the secretory pathway.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available