May Meeting

Speaker: Aleksandra Nita-Lazar, NIAID, NIH

Topic: Macrophage signaling mechanisms: deciphering protein networks

Date: Monday, May 15, 2023

Time: 6:15 pm Dinner, 7:15 pm Presentation

Location: Shimadzu Scientific Instrument, Inc. Training Center 7100 Riverwood Drive, Columbia, MD 21046 (Directions)
This will be an in-person meeting. Attendees are required to show a vaccine card (either at the door or in advance using the web form) . If you have submitted your vaccine card before, your status is already recorded.

Dinner: Please RSVP to Andy Qi (andy.yue.qi@gmail.com) by Friday, May 12 if you will be attending the dinner.

Abstract: Toll-like receptor (TLR) signaling in macrophages is essential for generating effective innate immune responses. Quantitative differences dependent on the dose and timing of the stimulus critically affect cell function and often involve proteins that are not components of widely shared transduction pathways. Mathematical modeling is an important approach to better understand how these signaling networks function in time and space. We have successfully modeled the S1P signaling pathway in macrophages using selected reaction monitoring (SRM) to measure the absolute abundance of the pathway proteins. The resulting values became parameters in a computational pathway model. To model the TLR signaling networks, we developed assays for the canonical
TLR signaling pathway and related proteins and phosphoproteins and used parallel reaction monitoring (PRM) with heavy-labeled internal peptide standards to quantify protein and phosphorylated protein molecule numbers per cell in untreated and LPS-stimulated macrophages. The absolute protein abundance values were entered into a model of the TLR pathway developed using Simmune, the rule-based modeling tool with a visual interface. To reach beyond basal level quantification, the TLR signaling network model is tested further and combined with global proteomic approaches to discover
biologically important new proteins, protein complexes and PTMs involved in this innate immune pathway. The protein and PTM levels are quantified in macrophages under diverse, but well-defined conditions (different TLR ligands, whole pathogens, and cells with mutations in specific signaling molecules). These data will allow to parameterize and test the TLR network model under a variety of conditions. Together, the interconnected projects will lead to the better understanding how the immune signaling pathways are regulated and activated during an infection. This research was supported by the Intramural Research Program of NIAID, NIH.

Thank you to our 2022-2023 sponsors!

April Meeting

Joint with the Washington Chromatography Discussion Group

Speaker: Ina Nemet, Lerner Research Institute, Cleveland Clinic

Topic: Dissecting gut microbial metabolic pathways in cardiometabolic diseases by mass spectrometry-based metabolic approaches

Date: Monday, April 17, 2023

Time: 6:15 pm Dinner, 7:15 pm Presentation

Location: Shimadzu Scientific Instrument, Inc. Training Center 7100 Riverwood Drive, Columbia, MD 21046 (Directions)
This will be an in-person meeting. Attendees are required to show a vaccine card (either at the door or in advance using the web form) . If you have submitted your vaccine card before, your status is already recorded.

Dinner: Please RSVP to Andy Qi (andy.yue.qi@gmail.com) by Friday, April 14 if you will be attending the dinner.

Abstract: Elucidating pathways that contribute to cardio-metabolic diseases beyond traditional risk factors, is important for developing new strategies effective in preventing and reducing disease progression. Multiple diseases including cardiovascular disease, obesity, type 2 diabetes, metabolic syndrome, and hypertension are associated with altered gut microbial structure and function. Rational design of personalized therapeutic strategies that target gut microbiota driven pathways require a comprehensive understanding of gut microbial metabolism and the relationship between systemic levels of metabolites and disease risks.
Recently we have clinically and mechanistically linked phenylacetylglutamine (PAGln), a gut microbially derived metabolite from aromatic amino acid phenylalanine, with cardiovascular disease risks. PAGln was elevated in diabetics and predicted incident risks for cardiovascular events independent of blood glucose levels. Mechanistic studies reveal PAGln acts via the adrenergic receptor. We have continued mapping out microbial metabolic pathways that are associated with adverse phenotypes in the host. The results of these studies will provide a meaningful starting point for the development of new therapeutic treatments for cardiometabolic diseases aimed at shifting nutrient fermentation away from metabolites associated with disease toward ones that are beneficial to the host.

Thank you to our 2022-2023 sponsors!

March Meeting

Speaker: Robert Cole, Johns Hopkins University School of Medicine

Topic: What’s on your albumin?

Date: Monday, March 20, 2023

Time: 6:15 pm Dinner, 7:15 pm Presentation

Location: Shimadzu Scientific Instrument, Inc. Training Center 7100 Riverwood Drive, Columbia, MD 21046 (Directions)
This will be an in-person meeting. Attendees are required to show a vaccine card (either at the door or in advance using the web form) . If you have submitted your vaccine card before, your status is already recorded.

Dinner: Please RSVP to Andy Qi (andy.yue.qi@gmail.com) by Friday, March 17 if you will be attending the dinner.

Abstract: Environmental exposures contribute to chronic disease risk substantially more than heritable genetic variants. Outdoor air pollution is a complex environmental
mixture which is responsible for over 4 million deaths each year, an impact that is projected to rise over the next several decades. The International Agency for Research on Cancer (IARC) has declared outdoor air pollution to be Group 1 human carcinogen. Outdoor air is a complex mixture of volatile organic toxins and carcinogens (e.g., aldehydes and benzene), sulfur dioxide, nitrogen oxides, polycyclic aromatic hydrocarbons, and particulate matter. Assessing personal exposure to environmental toxicants in complex mixtures, such as outdoor air pollution, is a critical challenge for predicting disease risk. Thus, developing and validating biomarkers which reveal exposure to these complex mixtures would advance individual risk analysis. Using human serum albumin (HSA)-based biomonitoring, we reported dosimetric relationships between adducts at HSA Cys34 and ambient air pollutant levels. However, modifications at other sites in HSA may reveal a great number of novel adducts and provide a panel of exposure biomarkers for disease risk. To bridge this gap, we developed a novel untargeted mass spectrometry-based method, termed Pan-Protein Adductomics (PPA),
to agnostically detect and quantify modifications at multiple residues in human serum albumin (HSA). Our PPA method combines nanoflow-liquid chromatography, gas-phase fractionation, overlapping-window data-independent acquisition, and high-resolution tandem mass spectrometry to assay modifications on HSA. Our initial application of PPA is to assess temporal changes in HSA modifications in non-smoking women exposed to high levels of outdoor air pollution. While we are currently focused on modifications in albumin, the PPA approach is applicable to any protein and may expand the knowledge base of protein modifications.

Thank you to our 2022-2023 sponsors!

Free Virtual Lipidomics Workshop

March 20-21, 2023, via Zoom

This fast-paced, interactive workshop will cover the fundamentals of lipid data acquisition, and data processing on a mass spectrometer. It is open to those who have at least 6 months of mass spec hands-on use and/or experience processing mass spec data.

This Zoom workshop will be a mix of lectures, demos and discussion, topics include:
Research Lecture by Alan Saghatelian of the Salk Institute for Biological Sciences
Chromatography and MS Acquisition Lecture
Method Setup on QExactive, Tribrid and Exploris Demo
Lipid Overview Lecture
Lipidex & Compound Discover Demo
Quality Control Lecture
Data Analysis Tools Lecture
Library Building Demo

APPLICATION CLOSES March 15 or when capacity is reached
Details can be found at https://www.ncqbcs.com/resources/training/lipidomics-training

LipidsWorkshopFlyer

February Meeting

Speaker: Zachary Goecker, NIST

Topic: Developing Reproducible Methods in Site-Specific Glycosylation Analysis

Date: Monday, February 27, 2023

Time: 6:15 pm Dinner, 7:15 pm Presentation

Location: Shimadzu Scientific Instrument, Inc. Training Center 7100 Riverwood Drive, Columbia, MD 21046 (Directions)
This will be an in-person meeting. Attendees are required to show a vaccine card (either at the door or in advance using the web form) . If you have submitted your vaccine card before, your status is already recorded.

Dinner: Please RSVP to Andy Qi (andy.yue.qi@gmail.com) by Friday, February 24 if you will be attending the dinner.

Abstract: Many proteins are glycosylated at multiple sites, and each site can contain complex
distributions of attached glycans. Until recently, reliable determinations of these distributions have not been possible. Instead, glycosylation analysis of glycoproteins has been primarily accomplished by releasing and identifying glycans, thereby losing all protein site-specific information. This glycomics approach works well for understanding the diversity of glycosylation in a digest, but does not answer questions relevant to site-specific structures (glycan microheterogeneity) and therefore is not as useful in determining direct functional implications. Recent developments in high resolution mass spectrometry have opened the door for the identification of intact glycopeptides. This study assesses methods in site-specific analysis of intact glycopeptides to demonstrate
reproducibility and measure variation in glycosylation quantitatively between different experimental factors. Here, we report the use of stepped HCD fragmentation, contingent ion scan, and many downstream data filters for the production of highly robust and reproducible glycosylation distribution spectra. In this presentation, we will report site-specific glycosylation profiles on glycoproteins from Sars-Cov-2, influenza, blood serum, and breastmilk. Results demonstrate that glycosylation profiles are highly reproducible among replicates and different digestion methods. However, glycosylation patterns do change based on factors such as source, proteins sequence, and glycosylation site.

Thank you to our 2022-2023 sponsors!