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14 pages, 3065 KiB

Open AccessArticle

The Introduction of Detergents in Thermal Proteome Profiling Requires Lowering the Applied Temperatures for Efficient Target Protein Identification

by Yuying Ye, Kejia Li, Yanni Ma, Xiaolei Zhang, Yanan Li, Ting Yu, Yan Wang and Mingliang Ye

Molecules 2023, 28(12), 4859; https://doi.org/10.3390/molecules28124859 - 20 Jun 2023

Cited by 2 | Viewed by 2412

Abstract

Although the use of detergents in thermal proteome profiling (TPP) has become a common practice to identify membrane protein targets in complex biological samples, surprisingly, there is no proteome-wide investigation into the impacts of detergent introduction on the target identification performance of TPP. [...] Read more.

Although the use of detergents in thermal proteome profiling (TPP) has become a common practice to identify membrane protein targets in complex biological samples, surprisingly, there is no proteome-wide investigation into the impacts of detergent introduction on the target identification performance of TPP. In this study, we assessed the target identification performance of TPP in the presence of a commonly used non-ionic detergent or a zwitterionic detergent using a pan-kinase inhibitor staurosporine, our results showed that the addition of either of these detergents significantly impaired the identification performance of TPP at the optimal temperature for soluble target protein identification. Further investigation showed that detergents destabilized the proteome and increased protein precipitation. By lowering the applied temperature point, the target identification performance of TPP with detergents is significantly improved and is comparable to that in the absence of detergents. Our findings provide valuable insight into how to select the appropriate temperature range when detergents are used in TPP. In addition, our results also suggest that the combination of detergent and heat may serve as a novel precipitation-inducing force that can be applied for target protein identification. Full article

(This article belongs to the Special Issue Advances and Future Perspective of Mass Spectrometry Analysis in Proteomics)

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23 pages, 3919 KiB

Open AccessArticle

Gene Ontology (GO)-Driven Inference of Candidate Proteomic Markers Associated with Muscle Atrophy Conditions

by Angelique Stalmach, Ines Boehm, Marco Fernandes, Alison Rutter, Richard J. E. Skipworth and Holger Husi

Molecules 2022, 27(17), 5514; https://doi.org/10.3390/molecules27175514 - 27 Aug 2022

Cited by 3 | Viewed by 3073

Abstract

Skeletal muscle homeostasis is essential for the maintenance of a healthy and active lifestyle. Imbalance in muscle homeostasis has significant consequences such as atrophy, loss of muscle mass, and progressive loss of functions. Aging-related muscle wasting, sarcopenia, and atrophy as a consequence of [...] Read more.

Skeletal muscle homeostasis is essential for the maintenance of a healthy and active lifestyle. Imbalance in muscle homeostasis has significant consequences such as atrophy, loss of muscle mass, and progressive loss of functions. Aging-related muscle wasting, sarcopenia, and atrophy as a consequence of disease, such as cachexia, reduce the quality of life, increase morbidity and result in an overall poor prognosis. Investigating the muscle proteome related to muscle atrophy diseases has a great potential for diagnostic medicine to identify (i) potential protein biomarkers, and (ii) biological processes and functions common or unique to muscle wasting, cachexia, sarcopenia, and aging alone. We conducted a meta-analysis using gene ontology (GO) analysis of 24 human proteomic studies using tissue samples (skeletal muscle and adipose biopsies) and/or biofluids (serum, plasma, urine). Whilst there were few similarities in protein directionality across studies, biological processes common to conditions were identified. Here we demonstrate that the GO analysis of published human proteomics data can identify processes not revealed by single studies. We recommend the integration of proteomics data from tissue samples and biofluids to yield a comprehensive overview of the human skeletal muscle proteome. This will facilitate the identification of biomarkers and potential pathways of muscle-wasting conditions for use in clinics. Full article

(This article belongs to the Special Issue Advances and Future Perspective of Mass Spectrometry Analysis in Proteomics)

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15 pages, 17370 KiB

Open AccessArticle

Prognostic Value of Molecular Intratumor Heterogeneity in Primary Oral Cancer and Its Lymph Node Metastases Assessed by Mass Spectrometry Imaging

by Agata Kurczyk, Marta Gawin, Piotr Paul, Ewa Chmielik, Tomasz Rutkowski, Monika Pietrowska and Piotr Widłak

Molecules 2022, 27(17), 5458; https://doi.org/10.3390/molecules27175458 - 25 Aug 2022

Cited by 3 | Viewed by 1857

Abstract

Different aspects of intra-tumor heterogeneity (ITH), which are associated with the development of cancer and its response to treatment, have postulated prognostic value. Here we searched for potential association between phenotypic ITH analyzed by mass spectrometry imaging (MSI) and prognosis of head and [...] Read more.

Different aspects of intra-tumor heterogeneity (ITH), which are associated with the development of cancer and its response to treatment, have postulated prognostic value. Here we searched for potential association between phenotypic ITH analyzed by mass spectrometry imaging (MSI) and prognosis of head and neck cancer. The study involved tissue specimens resected from 77 patients with locally advanced oral squamous cell carcinoma, including 37 patients where matched samples of primary tumor and synchronous lymph node metastases were analyzed. A 3-year follow-up was available for all patients which enabled their separation into two groups: with no evidence of disease (NED, n = 41) and with progressive disease (PD, n = 36). After on-tissue trypsin digestion, peptide maps of all cancer regions were segmented using an unsupervised approach to reveal their intrinsic heterogeneity. We found that intra-tumor similarity of spectra was higher in the PD group and diversity of clusters identified during image segmentation was higher in the NED group, which indicated a higher level of ITH in patients with more favorable outcomes. Signature of molecular components that correlated with long-term outcomes could be associated with proteins involved in the immune functions. Furthermore, a positive correlation between ITH and histopathological lymphocytic host response was observed. Hence, we proposed that a higher level of ITH revealed by MSI in cancers with a better prognosis could reflect the presence of heterotypic components of tumor microenvironment such as infiltrating immune cells enhancing the response to the treatment. Full article

(This article belongs to the Special Issue Advances and Future Perspective of Mass Spectrometry Analysis in Proteomics)

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15 pages, 3234 KiB

Open AccessArticle

MALDI-MSI: A Powerful Approach to Understand Primary Pancreatic Ductal Adenocarcinoma and Metastases

by Juliana Pereira Lopes Gonçalves, Christine Bollwein, Anna Melissa Schlitter, Mark Kriegsmann, Anne Jacob, Wilko Weichert and Kristina Schwamborn

Molecules 2022, 27(15), 4811; https://doi.org/10.3390/molecules27154811 - 27 Jul 2022

Cited by 9 | Viewed by 2779

Abstract

Cancer-related deaths are very commonly attributed to complications from metastases to neighboring as well as distant organs. Dissociate response in the treatment of pancreatic adenocarcinoma is one of the main causes of low treatment success and low survival rates. This behavior could not [...] Read more.

Cancer-related deaths are very commonly attributed to complications from metastases to neighboring as well as distant organs. Dissociate response in the treatment of pancreatic adenocarcinoma is one of the main causes of low treatment success and low survival rates. This behavior could not be explained by transcriptomics or genomics; however, differences in the composition at the protein level could be observed. We have characterized the proteomic composition of primary pancreatic adenocarcinoma and distant metastasis directly in human tissue samples, utilizing mass spectrometry imaging. The mass spectrometry data was used to train and validate machine learning models that could distinguish both tissue entities with an accuracy above 90%. Model validation on samples from another collection yielded a correct classification of both entities. Tentative identification of the discriminative molecular features showed that collagen fragments (COL1A1, COL1A2, and COL3A1) play a fundamental role in tumor development. From the analysis of the receiver operating characteristic, we could further advance some potential targets, such as histone and histone variations, that could provide a better understanding of tumor development, and consequently, more effective treatments. Full article

(This article belongs to the Special Issue Advances and Future Perspective of Mass Spectrometry Analysis in Proteomics)

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17 pages, 4212 KiB

Open AccessArticle

Mass Spectrometry-Based Peptide Profiling of Haemolymph from Pterostichus melas Exposed to Pendimethalin Herbicide

by Donatella Aiello, Anita Giglio, Federica Talarico, Maria Luigia Vommaro, Antonio Tagarelli and Anna Napoli

Molecules 2022, 27(14), 4645; https://doi.org/10.3390/molecules27144645 - 21 Jul 2022

Cited by 3 | Viewed by 2453

Abstract

Pendimethalin-based herbicides are used worldwide for pre-emergence selective control of annual grasses and weeds in croplands. The endurance of herbicides residues in the environment has an impact on the soil biodiversity and fertility, also affecting non-target species, including terrestrial invertebrates. Carabid beetles are [...] Read more.

Pendimethalin-based herbicides are used worldwide for pre-emergence selective control of annual grasses and weeds in croplands. The endurance of herbicides residues in the environment has an impact on the soil biodiversity and fertility, also affecting non-target species, including terrestrial invertebrates. Carabid beetles are known as natural pest control agents in the soil food web of agroecosystems, and feed on invertebrates and weed seeds. Here, a mass spectrometry untargeted profiling of haemolymph is used to investigate Pterostichus melas metabolic response after to pendimethalin-based herbicide exposure. Mass spectrometric data are examined with statistical approaches, such as principal component analysis, for possible correlation with biological effects. Those signals with high correlation are submitted to tandem mass spectrometry to identify the associated biomarker. The time course exposure showed many interesting findings, including a significant downregulation of related to immune and defense peptides (M-lycotoxin-Ls4a, Peptide hormone 1, Paralytic peptide 2, and Serine protease inhibitor 2). Overall, the observed peptide deregulations concur with the general mechanism of uptake and elimination of toxicants reported for Arthropods. Full article

(This article belongs to the Special Issue Advances and Future Perspective of Mass Spectrometry Analysis in Proteomics)

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17 pages, 2658 KiB

Open AccessArticle

Forensic Discrimination of Differentially Sourced Animal Blood Using a Bottom-Up Proteomics Based MALDI MS Approach

by Katie Kennedy, Laura Cole, Matthias Witt, Mark Sealey and Simona Francese

Molecules 2022, 27(7), 2039; https://doi.org/10.3390/molecules27072039 - 22 Mar 2022

Cited by 1 | Viewed by 2980

Abstract

Recently published work has reported the development and application of a bottom-up proteomic approach to distinguish between human and animal blood (down to animal species level), by rapid screening using Matrix Assisted Laser Desorption Ionisation Mass Spectrometry (MALDI MS). In that study, it [...] Read more.

Recently published work has reported the development and application of a bottom-up proteomic approach to distinguish between human and animal blood (down to animal species level), by rapid screening using Matrix Assisted Laser Desorption Ionisation Mass Spectrometry (MALDI MS). In that study, it was additionally observed that intravenous animal blood exhibits different spectral profiles from blood collected within the animal chest cavity as well as from the diluted blood collected within packets of meat. In this follow-up study we explored the resulting hypothesis that, depending on how blood is shed or collected, protein biomarker profiles vary to the extent of systematically permitting a distinction between possible sources of blood (for example, flesh wound versus packaged meat). This intelligence may be important in reconstructing the dynamics of the crime. The combination of statistical analysis and tandem mass spectrometry has yielded additional animal blood markers as well as confirming the ability to correctly determine the animal species from which blood derived, regardless of the retailer selling it (amongst the five investigated). These data confirm the initial hypothesis and demonstrate the opportunity for the proteomics-MALDI combined approach to provide additional intelligence to the investigation of violent crimes when examining blood evidence. Full article

(This article belongs to the Special Issue Advances and Future Perspective of Mass Spectrometry Analysis in Proteomics)

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19 pages, 2828 KiB

Open AccessArticle

Semi-Quantitative MALDI Measurements of Blood-Based Samples for Molecular Diagnostics

by Matthew A. Koc, Senait Asmellash, Patrick Norman, Steven Rightmyer, Joanna Roder, Robert W. Georgantas III and Heinrich Roder

Molecules 2022, 27(3), 997; https://doi.org/10.3390/molecules27030997 - 1 Feb 2022

Cited by 1 | Viewed by 2182

Abstract

Accurate and precise measurement of the relative protein content of blood-based samples using mass spectrometry is challenging due to the large number of circulating proteins and the dynamic range of their abundances. Traditional spectral processing methods often struggle with accurately detecting overlapping peaks [...] Read more.

Accurate and precise measurement of the relative protein content of blood-based samples using mass spectrometry is challenging due to the large number of circulating proteins and the dynamic range of their abundances. Traditional spectral processing methods often struggle with accurately detecting overlapping peaks that are observed in these samples. In this work, we develop a novel spectral processing algorithm that effectively detects over 1650 peaks with over 3.5 orders of magnitude in intensity in the 3 to 30 kD m/z range. The algorithm utilizes a convolution of the peak shape to enhance peak detection, and accurate peak fitting to provide highly reproducible relative abundance estimates for both isolated peaks and overlapping peaks. We demonstrate a substantial increase in the reproducibility of the measurements of relative protein abundance when comparing this processing method to a traditional processing method for sample sets run on multiple matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) instruments. By utilizing protein set enrichment analysis, we find a sizable increase in the number of features associated with biological processes compared to previously reported results. The new processing method could be very beneficial when developing high-performance molecular diagnostic tests in disease indications. Full article

(This article belongs to the Special Issue Advances and Future Perspective of Mass Spectrometry Analysis in Proteomics)

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22 pages, 3185 KiB

Open AccessArticle

Proteomic and Bioinformatic Analysis of Decellularized Pancreatic Extracellular Matrices

by Ming Hu, Huanjing Bi, Deana Moffat, Margaret Blystone, Lillian DeCostanza, Tchilabalo Alayi, Kaiming Ye, Yetrib Hathout and Sha Jin

Molecules 2021, 26(21), 6740; https://doi.org/10.3390/molecules26216740 - 8 Nov 2021

Cited by 11 | Viewed by 4641

Abstract

Tissue microenvironments are rich in signaling molecules. However, factors in the tissue matrix that can serve as tissue-specific cues for engineering pancreatic tissues have not been thoroughly identified. In this study, we performed a comprehensive proteomic analysis of porcine decellularized pancreatic extracellular matrix [...] Read more.

Tissue microenvironments are rich in signaling molecules. However, factors in the tissue matrix that can serve as tissue-specific cues for engineering pancreatic tissues have not been thoroughly identified. In this study, we performed a comprehensive proteomic analysis of porcine decellularized pancreatic extracellular matrix (dpECM). By profiling dpECM collected from subjects of different ages and genders, we showed that the detergent-free decellularization method developed in this study permits the preservation of approximately 62.4% more proteins than a detergent-based method. In addition, we demonstrated that dpECM prepared from young pigs contained approximately 68.5% more extracellular matrix proteins than those prepared from adult pigs. Furthermore, we categorized dpECM proteins by biological process, molecular function, and cellular component through gene ontology analysis. Our study results also suggested that the protein composition of dpECM is significantly different between male and female animals while a KEGG enrichment pathway analysis revealed that dpECM protein profiling varies significantly depending on age. This study provides the proteome of pancreatic decellularized ECM in different animal ages and genders, which will help identify the bioactive molecules that are pivotal in creating tissue-specific cues for engineering tissues in vitro. Full article

(This article belongs to the Special Issue Advances and Future Perspective of Mass Spectrometry Analysis in Proteomics)

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