Publications by authors named "Kyle Juetten"
Article Synopsis
- The main protease (M) of SARS-CoV-2 is crucial for the virus's maturation and is the target of the COVID-19 treatment Paxlovid, but there's a pressing need to find new inhibitors due to potential viral resistance.
- The study utilized advanced techniques like native mass spectrometry and UV photodissociation to analyze the structure of M and how it interacts with potential covalent inhibitors.
- Results indicated that certain inhibitors enhance the stability of M by creating dimeric complexes with higher melting temperatures and lower charge states, providing valuable insights into how these inhibitors work.
Owing to its ability to generate extensive fragmentation of proteins, ultraviolet photodissociation (UVPD) mass spectrometry (MS) has emerged as a versatile ion activation technique for the structural characterization of native proteins and protein complexes. Interpreting these fragmentation patterns provides insight into the secondary and tertiary structures of protein ions. However, the inherent complexity and diversity of proteins often pose challenges in resolving their numerous conformations.
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- - Charge detection mass spectrometry (CD-MS) allows for the analysis of large and complex analytes by examining individual ion signals, but it requires long analysis times due to the need for numerous scans to achieve reliable data.
- - The slow speed of CD-MS presents challenges for integration with fast techniques like chromatography, as limited scans may not provide adequate ion statistics from a single injection.
- - UniChromCD, a new module in the open-source UniDec package, simplifies the processing of CD-MS data by combining it with advanced demultiplexing methods, enhancing analysis and visualization of time-resolved data while supporting techniques like size-exclusion chromatography and ion mobility.
Article Synopsis
- Phosphatases play a crucial role in cellular processes by working with kinases through phosphorylation and dephosphorylation, making them important targets for drug development.! -
- The study employs ultraviolet photodissociation to investigate how two covalent inhibitors, T65 and rabeprazole, bind to the human SCP1 phosphatase and its mutant C181A, which has a nonreactive cysteine replaced.! -
- Top-down mass spectrometry analysis helps identify where these inhibitors attach to the proteins and assess the reactivity of different cysteine residues involved in the binding process.!
Mass-spectrometry-based methods have made significant progress in the characterization of post-translational modifications (PTMs) in peptides and proteins; however, room remains to improve fragmentation methods. Ideal MS/MS methods are expected to simultaneously provide extensive sequence information and localization of PTM sites and retain labile PTM groups. This collection of criteria is difficult to meet, and the various activation methods available today offer different capabilities.
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- RNA polymerase II (Pol II) undergoes important post-translational modifications on its C-terminal domain (CTD), which help regulate transcription by attracting different proteins at various stages of the transcription process.
- *The phosphorylation of specific serine residues (Ser5 and Ser2) on the CTD is crucial, as it occurs in relation to the transcription phases and influences which transcriptional regulators bind to Pol II.
- *The study identified calcium homeostasis endoplasmic reticulum protein (CHERP) as a significant regulatory protein that associates with the phosphorylated CTD, impacting alternative splicing when disrupted.
The impact of supercharging on the fragmentation patterns of six proteins, ubiquitin, cytochrome c, staph nuclease, myoglobin, dihydrofolate reductase, and carbonic anhydrase, was investigated for five activation methods, HCD, ETD, EThcD, 213 nm UVPD, and 193 nm UVPD under denaturing conditions. Changes in sequence coverage, alterations in the number and abundance of preferential cleavages (N-terminal to proline, C-terminal to aspartic or glutamic acid, adjacent to aromatic residues), and changes in individual fragment ion abundances were evaluated. Large decreases in sequence coverage were observed upon supercharging of proteins activated by HCD, whereas modest gains were observed for ETD.
View Article and Find Full Text PDFMS-TAFI: A Tool for the Analysis of Fragment Ions Generated from Intact Proteins.
J Proteome Res
February 2023
Tandem mass spectrometry (MS/MS) spectra of intact proteins can be difficult to interpret owing to the variety of fragment ion types and abundances. This information is crucial for maximizing the information derived from top-down mass spectrometry of proteins and protein complexes. MS-TAFI (Mass Spectrometry Tool for the Analysis of Fragment Ions) is a free Python-based program which offers a streamlined approach to the data analysis and visualization of deconvoluted MS/MS data of intact proteins.
View Article and Find Full Text PDFThe direct correlation between proteoforms and biological phenotype necessitates the exploration of mass spectrometry (MS)-based methods more suitable for proteoform detection and characterization. Here, we couple nano-hydrophobic interaction chromatography (nano-HIC) to ultraviolet photodissociation MS (UVPD-MS) for separation and characterization of intact proteins and proteoforms. High linearity, sensitivity, and sequence coverage are obtained with this method for a variety of proteins.
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