||Matthew B. Renfrow, Ph.D.
Assistant Professor, Department of Biochemistry and Molecular Genetics
Areas of Focus: Application of high resolution Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS) to answer specific biological questions.
The work in my lab centers on the application of high resolution Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS) to answer specific biological questions. There are two main areas of interest in the lab:
Ligand mediated RXR nuclear receptor transcriptional activation:
Retinoid X receptor (RXR) (Figure 1 top) is a ligand dependent nuclear receptor transcription factor fundamental in regulation of cellular proliferation, differentiation and growth. Small molecule binding repositions key structural elements in RXR ligand binding domain (LBD), presenting/burying coregulator interaction surfaces. Agonist binding removes corepressor proteins and promotes interactions with transcriptional coactivators. RXR associated transcription complexes are of therapeutic interest for prevention and treatment of breast cancer. We are interested in the process of how ligand binding results in a conformational change in the ligand binding domain of RXR nuclear receptor transcription factor that leads to a specific transcriptional response. This ligand dependent process is often studied by use of X-ray crystallography. However, many of these resulting structures are very similar. Our lab makes use of in-solution Hydrogen deuterium exchange (HD X) coupled with high resolution mass spectrometry (MS) (Figure 1 bottom) to compare and contrast the changes in in-solution conformational dynamics of the RXR LBD when bound to different ligands and coactivators. These HD X profiles provide a dynamic component to the structural analysis and can detect differences that ligand binding induces in the RXR LBD that are not observed in x-ray crystal analysis. These dynamic changes are quantative and can be used as structural markers to screen novel rexinoids for similar binding characteristics. We also correlate our results with more traditional molecular biology assays for characterizing novel RXR ligands in terms of ligand affinity and RXR activation. Overall, our goal is to provide a unique set of tools and perspective for understanding the process of ligand induced nuclear receptor activation.
Analysis of clustered sites of O-glycosylation:
Glycosylation is one of the most common post-translational modifications of proteins. It is estimated that over half of mammalian proteins are glycosyalted. Several autoimmune disorders and chronic inflammatory diseases exhibit abnormal glycosylation of serum immunoglobulins. A variety of proteins are postranslationally modified with clustered sites of O-glycosylation. Serine and Threonine rich stretches within the amino acid sequence that have short O-glycan chains. Examples include the immunoglobulin A (A1 isotype), mucins, and bacterial cell surface proteins. For a given protein with sites of clustered O-glycans, the protein isolated from a single source is a population of variably O-glycosylated isoforms that usually show a distinct distribution of microheterogeneity in terms of number of chains, the sites of attachment and O-glycan composition at a given amino acid. Characterizing these clustered sites and understanding how the distributions change under differenct biological conditions or disease states is an analytical challenge. IgA nephropathy (IgAN) is the most common primary glomerulonephritis worldwide, with about 20-40% of patients developing end-stage renal failure. It is characterized by mesangial deposits of IgA1-containing immune complexes. The carbohydrate side chains of IgA1 molecules play a pivotal role in the pathogenesis of IgAN 5-13. IgA1 contains a hinge region between the first and second heavy chain constant region domains with a high content of proline, serine, and threonine and usually have three to five O-linked glycan chains (Figure 2, middle). Our group makes use of high resolution FT-ICR MS to provide an accurate profile of entire population of O-glycosylated IgA1 proteins to identify the pathogenic forms contributing to the pathogenesis of IgAN. This includes the site-specific localization and characterization of individual O-glycan chains by use of electron capture/transfer dissociation (ECD & ETD)(Figure 2 bottom). Our goal is to identify the aberrant IgA1 O-glycosylation pattern that leads to the mesangial deposits of IgA1-containing immune complexes. This may lead to alternative methods for diagnosing and monitoring the disease as well as identifying targets for therapeutic intervention.
Dr. Matthew Renfrow is an Assistant Professor in the Department of Biochemistry and Molecular Genetics. Dr. Renfrow received a B.S. degree in both Biochemistry and Recombinant Genetics from Western Kentucky University in 1996. He received his Ph.D. degree from the University of Georgia in 2002. His postdoctoral studies were completed at National High Magnetic Field Laboratory, Florida State University in 2004. Dr. Renfrow joined the faculty as an Assistant Professor at UAB in 2004. Please see websites for research information: www.uab.edu/BiomedFTICR/ and www.uab.edu/BiomedFTICR/renfrow/.