Culture in Transit: Digitizing and Democratizing NYC’s Cultural Heritage

Presentation from the MARAC conference in Pittsburgh, PA on April 14–16, 2016. S4 - Culture in Transit: Digitizing and Democratizing NYC’s Cultural Heritage.

Analysis of Federal Policy on Public Access to Scientific Research Data

Presentation from the MARAC conference in Pittsburgh, PA on April 14–16, 2016. S13 - Student Poster Session; Analysis of Federal Policy on Public Access to Scientific Research Data

Grant Cycles, Deadlines, and Labor Advocacy: The Changing Work of Project Archivists

Presentation from the MARAC conference in Pittsburgh, PA on April 14–16, 2016. S7 - Grant Cycles, Deadlines, and Labor Advocacy: The Changing Work of Project Archivists.

How to Create an Oral History Program

Presentation from the MARAC conference in Pittsburgh, PA on April 14–16, 2016. S3 - How to Create an Oral History Program.

PA Photos & Documents: Exploring Pennsylvania’s digitized documents and photographs

Presentation from the MARAC conference in Pittsburgh, PA on April 14–16, 2016. S24; - Pittsburgh Pop-Up #2.

The Duchamp Research Portal: Moving an Idea to Proof of Concept

Presentation from the MARAC conference in Pittsburgh, PA on April 14–16, 2016. S15 - The Duchamp Research Portal: Moving an Idea to Proof of Concept.

In the Beginning: Building an Archive from the Ground Up

Presentation from the MARAC conference in Pittsburgh, PA on April 14–16, 2016. S23 - In the Beginning: Building an Archive from the Ground Up

Communities in Collaboration: Building the Empire Archival Discovery Cooperative

Presentation from the MARAC conference in Pittsburgh, PA on April 14–16, 2016. S24; - Pittsburgh Pop-Up #2.

Pittsburgh and Philadelphia Can Work Together: Processing the Arlen Specter Collection at the University of Pittsburgh

Presentation from the MARAC conference in Pittsburgh, PA on April 14–16, 2016. S2 - Making Friends: The Highs, Lows, and Challenges of Inter-Repository Archival Relationships

CHARACTERIZATION OF POLYUBIQUITIN CHAINS BY MASS SPECTROMETRY

The work outlined in this dissertation will allow biochemists and cellular biologists to characterize polyubiquitin chains involved in their cellular environment by following a facile mass spectrometric based workflow. The characterization of polyubiquitin chains has been of interest since their discovery in 1984. The profound effects of ubiquitination on the movement and processing of cellular proteins depend exclusively on the structures of mono and polyubiquitin modifications anchored or unanchored on the protein within the cellular environment. However, structure-function studies have been hindered by the difficulty in identifying complex chain structures due to limited instrument capabilities of the past. Genetic mutations or reiterative immunoprecipitations have been used previously to characterize the polyubiquitin chains, but their tedium makes it difficult to study a broad ubiquitinome. Top-down and middle-out mass spectral based proteomic studies have been reported for polyubiquitin and have had success in characterizing parts of the chain, but no method to date has been successful at differentiating all theoretical ubiquitin chain isomers (ubiquitin chain lengths from dimer to tetramer alone have 1340 possible isomers). The workflow presented here can identify chain length, topology and linkages present using a chromatographic-time-scale compatible, LC-MS/MS based workflow. To accomplish this feat, the strategy had to exploit the most recent advances in top-down mass spectrometry. This included the most advanced electron transfer dissociation (ETD) activation and sensitivity for large masses from the orbitrap Fusion Lumos. The spectral interpretation had to be done manually with the aid of a graphical interface to assign mass shifts because of a lack of software capable to interpret fragmentation across isopeptide linkages. However, the method outlined can be applied to any mass spectral based system granted it results in extensive fragmentation across the polyubiquitin chain; making this method adaptable to future advances in the field.