Biology subjects

Pergande, M. R.

Publications and source records attributed to Pergande, M. R..

5 recordsLinked to original sources

bioRxiv2023

Comprehensive Metabolomic Analysis of Human Heart Tissue Enabled by Parallel Metabolite Extraction and High-Resolution Mass Spectrometry

The heart contracts incessantly and requires a constant supply of energy, utilizing numerous metabolic substrates such as fatty acids, carbohydrates, lipids, and amino acids to supply its high energy demands. Therefore, a comprehensive analysis of various metabolites is urgently needed for understanding cardiac metabolism; however, complete metabolome analyses remain challenging due to the broad range of metabolite polarities which makes extraction and detection difficult. Herein, we implemented parallel metabolite extractions and high-resolution mass spectrometry (MS)-based methods to obtain a comprehensive analysis of the human heart metabolome. To capture the diverse range of metabolite polarities, we first performed six parallel liquid-liquid extractions (three monophasic, two biphasic, and one triphasic extractions) of healthy human donor heart tissue. Next, we utilized two complementary MS platforms for metabolite detection - direct-infusion ultrahigh-resolution Fourier-transform ion cyclotron resonance (DI-FTICR) and high-resolution liquid chromatography quadrupole time-of-flight tandem MS (LC-Q-TOF MS/MS). Using DI-FTICR MS, 9,521 metabolic features were detected where 7,699 were assigned a chemical formula and 1,756 were assigned an annotated by accurate mass assignment. Using LC-Q-TOF MS/MS, 21,428 metabolic features were detected where 626 metabolites were identified based on fragmentation matching against publicly available libraries. Collectively, 2276 heart metabolites were identified in this study which span a wide range of polarities including polar (benzenoids, alkaloids and derivatives and nucleosides) as well as non-polar (phosphatidylcholines, acylcarnitines, and fatty acids) compounds. The results of this study will provide critical knowledge regarding the selection of appropriate extraction and MS detection methods for the analysis of the diverse classes of human heart metabolites. Table of Contents Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=108 SRC="FIGDIR/small/558013v1_ufig1.gif" ALT="Figure 1"> View larger version (27K): org.highwire.dtl.DTLVardef@1db91c8org.highwire.dtl.DTLVardef@400d81org.highwire.dtl.DTLVardef@136b221org.highwire.dtl.DTLVardef@1e8d3b9_HPS_FORMAT_FIGEXP M_FIG C_FIG

molecular biology↗

bioRxiv2023

High-throughput Extracellular Matrix Proteomics of Human Lungs Enabled by Photocleavable Surfactant and diaPASEF

The extracellular matrix (ECM) is a complex assembly of proteins that provide interstitial scaffolding and elastic recoil to human lungs. The pulmonary extracellular matrix (ECM) is increasingly recognized as an independent bioactive entity by creating biochemical and mechanical signals that influence disease pathogenesis, making it an attractive therapeutic target. However, the pulmonary ECM proteome ("matrisome") remains challenging to analyze by mass spectrometry due to its inherent biophysical properties and relatively low abundance. Here, we introduce a strategy designed for rapid and efficient characterization of the human pulmonary ECM using the photocleavable surfactant Azo. We coupled this approach with trapped ion mobility MS with diaPASEF to maximize depth of matrisome coverage. Using this strategy, we identify nearly 400 unique matrisome proteins with excellent reproducibility that are known to be important in lung biology, including key insoluble ECM proteins.

systems biology↗

bioRxiv2023

Mass spectrometry-based multi-omics identifies metabolic signatures of sarcopenia in rhesus monkey skeletal muscle

Sarcopenia is a progressive disorder characterized by age-related loss of skeletal muscle mass and function. Although significant progress has been made over the years to identify the molecular determinants of sarcopenia, the precise mechanisms underlying the age-related loss of contractile function remains unclear. Advances in omics technologies, including mass spectrometry-based proteomic and metabolomic analyses, offer great opportunities to better understand sarcopenia. Herein, we performed mass spectrometry-based analyses of the vastus lateralis from young, middle-aged, and older rhesus monkeys to identify molecular signatures of sarcopenia. In our proteomic analysis, we identified numerous proteins that change with age, including those involved in adenosine triphosphate and adenosine monophosphate metabolism as well as fatty acid beta oxidation. In our untargeted metabolomic analysis, we identified multiple metabolites that changed with age largely related to energy metabolism including fatty acid beta oxidation. Pathway analysis of age-responsive proteins and metabolites revealed changes in muscle structure and contraction as well as lipid, carbohydrate, and purine metabolism. Together, this study discovers new metabolic signatures and offer new insights into the molecular mechanism underlying sarcopenia for the evaluation and monitoring of therapeutic treatment of sarcopenia. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=67 SRC="FIGDIR/small/551315v1_ufig1.gif" ALT="Figure 1"> View larger version (15K): org.highwire.dtl.DTLVardef@1dc0addorg.highwire.dtl.DTLVardef@14d73b8org.highwire.dtl.DTLVardef@1e8733forg.highwire.dtl.DTLVardef@1e65408_HPS_FORMAT_FIGEXP M_FIG C_FIG Graphical Abstract

biochemistry↗

bioRxiv2023

DBDA matrix increases ion abundance of fatty acids and sulfatides in MALDI-TOF and mass spectrometry imaging studies

MALDI-TOF MS is a powerful tool to analyze biomolecules owing to its soft ionization nature and generally results in simple spectra of singly charged ions. Moreover, implementation of the technology in imaging mode provides a means to spatially map analytes in situ. Recently, a new matrix, DBDA (N1,N4-dibenzylidenebenzene-1,4-diamine) was reported to facilitate the ionization of free fatty acids in the negative ion mode. Building on this finding, we sought to implement DBDA for MALDI mass spectrometry imaging studies in brain tissue and successfully map oleic acid, palmitic acid, stearic acid, docosahexaenoic acid and arachidonic acid using mouse brain sections. Moreover, we hypothesized that DBDA would provide superior ionization for sulfatides, a class of sulfolipids, with multiple biological functions. Herein we also demonstrate that DBDA is ideal for MALDI mass spectrometry imaging of fatty acids and sulfatides in brain tissue sections. Additionally, we show enhanced ionization of sulfatides using DBDA compared to three different traditionally used MALDI matrices. Together these results provide new opportunities for studies to measure sulfatides by MALDI-TOF MS including in imaging modes.

biochemistry↗

bioRxiv2023

MASH Native: A Unified Solution for Native Top-Down Proteomics Data Processing

Native top-down proteomics (nTDP) integrates native mass spectrometry (nMS) with top-down proteomics (TDP) to provide comprehensive analysis of protein complexes together with proteoform identification and characterization. Despite significant advances in nMS and TDP software developments, a unified and user-friendly software package for analysis of nTDP data remains lacking. Herein, we have developed MASH Native to provide a unified solution for nTDP to process complex datasets with database searching capabilities in a user-friendly interface. MASH Native supports various data formats and incorporates multiple options for deconvolution, database searching, and spectral summing to provide a one-stop shop for characterizing both native protein complexes and proteoforms. The MASH Native app, video tutorials, written tutorials and additional documentation are freely available for download at https://labs.wisc.edu/gelab/MASH_Explorer/MASHNativeSoftware.php. All data files shown in user tutorials are included with the MASH Native software in the download .zip file.

bioinformatics↗