Biomedical Research Education & Training
Faculty Member

Ohi, Melanie D., Ph.D
Associate Professor of Cell and Developmental Biology

Lab Url: http://www.mc.vanderbilt.edu/vumcdept/cellbio/ohimdlab

Phone Number: 936-7780

Email Address: melanie.ohi@vanderbilt.edu

Ohi, Melanie's picture
Academic history
B.S., Pacific Lutheran University
Ph.D., Vanderbilt
post-doctoral fellowship, Harvard Medical School

Office Address   Mailing Address

Office: 4160A MRBIII

4160 MRB III 37232


Research Keywords
Single particle electron microscopy, Cryo-electron microscopy, Spliceosome, Schizosaccharomyces pombe, bacterial toxins, pathogenesis,Biochemistry,Gene regulation,Infectious disease,Microbiology,Protein Structure,Structural Biology,Yeast

Research Specialty
Studying the structure and function of macromolecular machines

Research Description
Proteins carry out most cellular processes as members of dynamic multi-protein assemblies. Although progress has been made cataloging the constituents of specific complexes, we have only limited knowledge of how proteins assemble into macromolecular machines and how these machines perform their cellular functions. We are using yeast genetics, biochemistry and single particle cryo-electron microscopy (EM) to explore the structural and functional organization of complexes involved in pre-mRNA splicing and complexes involved in the protein ubiquitination pathway.

Single Particle Cryo-Electron Microscopy
Single particle cryo-EM is a powerful technique for determining the structures of large, dynamic complexes that are too difficult to crystallize. In this structural approach, purified complexes are applied to grids covered with holey carbon film and quickly frozen by plunging the grids into liquid ethane. The rapid freezing prevents water from forming ice crystals and embeds the molecules in a layer of vitrified (or amorphous) ice preserving the specimen in a near-native environment. Images of the preserved particles are taken using an electron microscope. Digital image processing methods are then used to produce 3D models from the images of complex particles trapped in vitrified ice.

The Spliceosome
Although the human genome contains ~25,000 genes, it is estimated that we make over 90,000 proteins. The disparity between our genome and our proteome can be explained by the activity of the spliceosome, a large macromolecular machine composed of RNA and protein components. This complex catalyzes the excision of non-coding introns from a pre-messenger RNA (pre-mRNA) to create a mature message (mRNA). Although the composition of the spliceosome is known, it remains a mystery how this dynamic machine functions. To understand spliceosome function and regulation it will be essential to develop three-dimensional (3D) pictures of how the numerous spliceosomal proteins and RNA components organize into one machine. A number of projects in the lab focus on the functional and structural characterization of stable spliceosomal complexes isolated from the fission yeast S. pombe.

Pore Forming Toxins
There are a number of projects in the lab that focus on studying the structure and function of pore forming bacterial toxins. We work with a multidisciplinary team of researchers to understand how these toxins function as both soluble and membrane inserted proteins, with the main goal of gaining a mechanistic understanding of how these toxins contribute to pathogenesis.

Publications
Archuleta, TL, Du, Y, English, CA, Lory, S, Lesser, C, Ohi, MD, Ohi, R, Spiller, BW. The Chlamydia effector, chlamydial outer protein N CopN), sequesters tubulin and prevents microtubule assembly. J Biol Chem, 2011

McLean, JR, Chaix, D, Ohi, MD, Gould, KL. State of the APC/C: organization, function, and structure. Crit Rev Biochem Mol Biol, 46(2), 118-36, 2011

Gonz?!lez-Rivera, C, Gangwer, KA, McClain, MS, Eli, IM, Chambers, MG, Ohi, MD, Lacy, DB, Cover, TL. Reconstitution of Helicobacter pylori VacA toxin from purified components. Biochemistry, 49(27), 5743-52, 2010

Pruitt, RN, Chambers, MG, Ng, KK, Ohi, MD, Lacy, DB. Structural organization of the functional domains of Clostridium difficile toxins A and B. Proc Natl Acad Sci U S A, 2010

Roberts-Galbraith, RH, Ohi, MD, Ballif, BA, Chen, JS, McLeod, I, McDonald, WH, Gygi, SP, Yates, JR, Gould, KL. Dephosphorylation of F-BAR protein Cdc15 modulates its conformation and stimulates its scaffolding activity at the cell division site. Mol Cell, 39(1), 86-99, 2010

Vander Kooi, CW, Ren, L, Xu, P, Ohi, MD, Gould, KL, Chazin, WJ. The Prp19 WD40 domain contains a conserved protein interaction region essential for its function. Structure, 18(5), 584-93, 2010

Huang, DT, Hunt, HW, Zhuang, M, Ohi, MD, Holton, JM, Schulman, BA. Basis for a ubiquitin-like protein thioester switch toggling E1-E2 affinity. Nature, 445(7126), 394-8, 2007

Ohi, MD, Feoktistova, A, Ren, L, Yip, C, Cheng, Y, Chen, JS, Yoon, HJ, Wall, JS, Huang, Z, Penczek, PA, Gould, KL, Walz, T. Structural organization of the anaphase-promoting complex bound to the mitotic activator Slp1. Mol Cell, 28(5), 871-85, 2007 PMCID:2197158

Ohi, MD, Ren, L, Wall, JS, Gould, KL, Walz, T. Structural characterization of the fission yeast U5.U2/U6 spliceosome complex. Proc Natl Acad Sci U S A, 104(9), 3195-200, 2007 PMCID:1805518

Crampton, D.J., Ohi, M.D., Qimron,E., Lee,S.J., Walz, T., and Richardson, C.. Oligomeric states of bacteriophage T7 gene 4 primase/helicase. Journal of Molecular Biology, 360, 667-677, 2006

Vander Kooi, CW, Ohi, MD, Rosenberg, JA, Oldham, ML, Newcomer, ME, Gould, KL, Chazin, WJ. The Prp19 U-box crystal structure suggests a common dimeric architecture for a class of oligomeric E3 ubiquitin ligases. Biochemistry, 45(1), 121-30, 2006 PMCID:2570371

Ohi, MD, Vander Kooi, CW, Rosenberg, JA, Ren, L, Hirsch, JP, Chazin, WJ, Walz, T, Gould, KL. Structural and functional analysis of essential pre-mRNA splicing factor Prp19p. Mol Cell Biol, 25(1), 451-60, 2005 PMCID:538785

Ohi, M.D., Li, Y., Cheng, Y. and Walz, T.. Negative staining and image classification-powerful tools in modern electron microscopy. Biol. Proc. Online, 6(1), 23-34, 2004

Wilkinson, CR, Dittmar, GA, Ohi, MD, Uetz, P, Jones, N, Finley, D. Ubiquitin-like protein Hub1 is required for pre-mRNA splicing and localization of an essential splicing factor in fission yeast. Curr Biol, 14(24), 2283-8, 2004

Ohi, MD, Vander Kooi, CW, Rosenberg, JA, Chazin, WJ, Gould, KL. Structural insights into the U-box, a domain associated with multi-ubiquitination. Nat Struct Biol, 10(4), 250-5, 2003

Ohi, MD, Gould, KL. Characterization of interactions among the Cef1p-Prp19p-associated splicing complex. RNA, 8(6), 798-815, 2002 PMCID:1370298

Ohi, MD, Link, AJ, Ren, L, Jennings, JL, McDonald, WH, Gould, KL. Proteomics analysis reveals stable multiprotein complexes in both fission and budding yeasts containing Myb-related Cdc5p/Cef1p, novel pre-mRNA splicing factors, and snRNAs. Mol Cell Biol, 22(7), 2011-24, 2002 PMCID:133674

Berry, LD, Feoktistova, A, Wright, MD, Gould, KL. The schizosaccharomyces pombe dim1(+) gene interacts with the anaphase-promoting complex or cyclosome (APC/C) component lid1(+) and is required for APC/C function. Mol Cell Biol, 19(4), 2535-46, 1999 PMCID:84046


Postdoctoral Position Available
N/A

Postdoctoral Position Details
N/A

Updated Date
06/12/2012