Center for Proteomics and Bioinformatics:
Department of Pharmacology:
Department of Ophthalmology and Visual Sciences:
|1997||Ph.D., Biochemistry, Osaka University|
|1998-2000||Post-doctoral Fellow, Biochemistry, Cleveland Clinic Foundation|
|1993-1997||Takara Shuzo Co., Ltd., Kusatsu, Japan|
|2000-2002||Project Scientist, Department of Ophthalmic Research, Cole Eye Institute, Cleveland Clinic Foundation, Cleveland, Ohio|
|2002-2006||Director, Proteomics/Mass Spectrometry Core Facility, University of North Dakota School of Medicine and Health Sciences, Grand Forks, North Dakota|
|2002-2006||Assistant Professor, Department of Biochemistry and Molecular Biology, University of North Dakota School of Medicine and Health Sciences, Grand Forks, North Dakota|
|2006-2015||Assistant Professor, Center for Proteomics & Bioinformatics, Case Western Reserve University, Cleveland, Ohio||2015-Present||Associate Professor, Center for Proteomics & Bioinformatics, Case Western Reserve University, Cleveland, Ohio|
1) Development of Analytical Methods
We are motivated to develop analytical methods for proteome profiling and protein structural characterization. Mass spectrometry is the pre-eminent technology in analytical protein chemistry. However, current methods on the front end of mass spectrometry analysis are not always taking advantage of modern mass spectrometry technologies. Development of advanced analytical methods on the front end of mass spectrometry analysis is therefore important in advancing the study of proteins. The following methods have been developed or are being developed in our laboratory.
Proteolytic 18O labeling
The proteolytic 18O labeling method is used to determine the relative expression levels of individual proteins between two samples. This technique utilizes a protease and water (H216O and H218O) to produce labeled peptides; peptides in one sample are labeled with 16O, and peptides in the other sample with 18O. Both samples are mixed in a 1:1 ratio and subjected to mass spectrometric analysis to identify and quantify the proteins from which the peptides originated. Our laboratory has contributed to establish highly efficient oxygen labeling conditions. Our current efforts are focused on establishing an experimental workflow that can be done entirely in a single tube in order to eliminate the peptide loss during the labeling procedure. This technique became one of the commonly used quantitative proteomics techniques in the field.
Histidine Hydrogen-Deuterium Exchange Mass Spectrometry (His-HDX-MS) measures the rates of HDX reaction of the imidazole C2-hydrogen in proteins using mass spectrometry. His-HDX-MS has been successfully applied to probe the structural changes of several proteins. These studies demonstrated that the HDX rates are extremely sensitive to the changes of electrostatic environment and solvent accessibilities of individual histidine residues. Our current efforts are focused on establishing a robust experimental platform for analyzing complex protein mixtures, which allows us to study proteins in a more biologically relevant context.
We have made heavy lysine-labeled SILAC-C. elegans by collaborating with Dr. Feng (Case Western Reserve University). This quantitative proteomics platform can be used for a multitude of biological studies using C. elegans. Our current efforts are focused on developing a method to measure the turnover of individual proteins in C. elegans using this platform.
2) Proteomic Projects
Mechanisms of diabetic retinopathy
Diabetic retinopathy is the most common diabetic eye disease and a leading cause of blindness in American adults. To date, however, no treatment that inhibits the retinopathy in patients is available. Interestingly, modulation of lysine acetylation events in diabetic animals by either inhibiting histone acetyltransferase (p300) or by activating NAD+-dependent protein deacetylase (Sirt1) has been shown to have beneficial effect on the development of retinopathy. This project aims to identify lysine acetylation sites and biological processes/pathways that are altered in diabetic retinopathy using cutting-edge mass spectrometry based quantitative proteomic techniques, in conjunction with an affinity enrichment technique for acetyl lysine containing peptides.
Mechanisms of aging using C. elegans
Elucidating the basic biochemical, genetic, and physiological processes underlying the progression of aging is essential to extend the healthy and active years of life. A numerous biochemical and genomic studies have been done on C. elegans to better understand the mechanisms of aging. However, knowledge of protein turnover in the global proteome and its relation to aging is lacking despite the fact that controlled and selective synthesis/degradation of proteins is known to be critical for most cellular processes. This project aims to reveal how the balance between protein synthesis and degradation changes over the life span of wild-type C. elegans at the individual protein level using the SILAC-C. elegans platform.