Mass spectrometry reveals the missing links in the assembly pathway of the bacterial 20 S proteasome.
The 20 S proteasome is an essential proteolytic particle, responsible for degrading short-lived and abnormal intracellular proteins. The 700-kDa assembly is comprised of 14 alpha-type and 14 beta-type subunits, which form a cylindrical architecture composed of four stacked heptameric rings (alpha7be...
| Main Authors: | , , , , |
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| Format: | Journal article |
| Language: | English |
| Published: |
2007
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| _version_ | 1797058836327038976 |
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| author | Sharon, M Witt, S Glasmacher, E Baumeister, W Robinson, C |
| author_facet | Sharon, M Witt, S Glasmacher, E Baumeister, W Robinson, C |
| author_sort | Sharon, M |
| collection | OXFORD |
| description | The 20 S proteasome is an essential proteolytic particle, responsible for degrading short-lived and abnormal intracellular proteins. The 700-kDa assembly is comprised of 14 alpha-type and 14 beta-type subunits, which form a cylindrical architecture composed of four stacked heptameric rings (alpha7beta7beta7alpha7). The formation of the 20 S proteasome is a complex process that involves a cascade of folding, assembly, and processing events. To date, the understanding of the assembly pathway is incomplete due to the experimental challenges of capturing short-lived intermediates. In this study, we have applied a real-time mass spectrometry approach to capture transient species along the assembly pathway of the 20 S proteasome from Rhodococcus erythropolis. In the course of assembly, we observed formation of an early alpha/beta-heterodimer as well as an unprocessed half-proteasome particle. Formation of mature holoproteasomes occurred in concert with the disappearance of half-proteasomes. We also analyzed the beta-subunits before and during assembly and reveal that those with longer propeptides are incorporated into half- and full proteasomes more rapidly than those that are heavily truncated. To characterize the preholoproteasome, formed by docking of two unprocessed half-proteasomes and not observed during assembly of wild type subunits, we trapped this intermediate using a beta-subunit mutational variant. In summary, this study provides evidence for transient intermediates in the assembly pathway and reveals detailed insight into the cleavage sites of the propeptide. |
| first_indexed | 2024-03-06T19:55:59Z |
| format | Journal article |
| id | oxford-uuid:259b31bf-af76-4ae7-837b-bf27caa64832 |
| institution | University of Oxford |
| language | English |
| last_indexed | 2024-03-06T19:55:59Z |
| publishDate | 2007 |
| record_format | dspace |
| spelling | oxford-uuid:259b31bf-af76-4ae7-837b-bf27caa648322022-03-26T11:56:27ZMass spectrometry reveals the missing links in the assembly pathway of the bacterial 20 S proteasome.Journal articlehttp://purl.org/coar/resource_type/c_dcae04bcuuid:259b31bf-af76-4ae7-837b-bf27caa64832EnglishSymplectic Elements at Oxford2007Sharon, MWitt, SGlasmacher, EBaumeister, WRobinson, CThe 20 S proteasome is an essential proteolytic particle, responsible for degrading short-lived and abnormal intracellular proteins. The 700-kDa assembly is comprised of 14 alpha-type and 14 beta-type subunits, which form a cylindrical architecture composed of four stacked heptameric rings (alpha7beta7beta7alpha7). The formation of the 20 S proteasome is a complex process that involves a cascade of folding, assembly, and processing events. To date, the understanding of the assembly pathway is incomplete due to the experimental challenges of capturing short-lived intermediates. In this study, we have applied a real-time mass spectrometry approach to capture transient species along the assembly pathway of the 20 S proteasome from Rhodococcus erythropolis. In the course of assembly, we observed formation of an early alpha/beta-heterodimer as well as an unprocessed half-proteasome particle. Formation of mature holoproteasomes occurred in concert with the disappearance of half-proteasomes. We also analyzed the beta-subunits before and during assembly and reveal that those with longer propeptides are incorporated into half- and full proteasomes more rapidly than those that are heavily truncated. To characterize the preholoproteasome, formed by docking of two unprocessed half-proteasomes and not observed during assembly of wild type subunits, we trapped this intermediate using a beta-subunit mutational variant. In summary, this study provides evidence for transient intermediates in the assembly pathway and reveals detailed insight into the cleavage sites of the propeptide. |
| spellingShingle | Sharon, M Witt, S Glasmacher, E Baumeister, W Robinson, C Mass spectrometry reveals the missing links in the assembly pathway of the bacterial 20 S proteasome. |
| title | Mass spectrometry reveals the missing links in the assembly pathway of the bacterial 20 S proteasome. |
| title_full | Mass spectrometry reveals the missing links in the assembly pathway of the bacterial 20 S proteasome. |
| title_fullStr | Mass spectrometry reveals the missing links in the assembly pathway of the bacterial 20 S proteasome. |
| title_full_unstemmed | Mass spectrometry reveals the missing links in the assembly pathway of the bacterial 20 S proteasome. |
| title_short | Mass spectrometry reveals the missing links in the assembly pathway of the bacterial 20 S proteasome. |
| title_sort | mass spectrometry reveals the missing links in the assembly pathway of the bacterial 20 s proteasome |
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