The terminal stage of ATP synthesis in the mitochondria of animal and human cells takes place within the inner membrane and requires 3 separate participants, the ATP synthase (F0F1). a double motor enzyme, the phosphate carrier (PIC), and the adenine nucleotide carrier (ANQ. During the past progress period that led to the discovery of several novel mechanistic and structural features of the mitochondrial ATP synthase (PNAS, 1998; JBC, 1999; JBC, 2000; Proteins, 2003), we discovered also that the above proteins exist as a single ATP synthase/PIC/ANC complex (> 0.7 million Da) that we named the """"""""ATP synthasome"""""""" (JBC, 2003). In addition to purifying this 17 subunit type complex, and identifying detergents that maintain it in a soluble, stable, dispersed form, we obtained a 3-D structure at 2.3 nm resolution by reconstructing negatively stained images of single molecules (JBC, 2004). Most recently, we have shown that the ATP svnthasome forms 2-dimensional (2D) crystalline sheets and is a functionally relevant complex as oligomycin. the classical inhibitor of mitochondrial oxidative phosphorylation. only inhibits the ATP svnthase when PIC and ANC are bound. Finally, in a project related to the primary mission of the NCI who funded this work, we found that the antimetabolite 3-bromopyruvic acid (3-BrPA) targets the ATP synthasome, rapidly and completely depletes cell ATP, and eradicates advanced stage cancers in all 19 treated cases in a rodent model (BBRC, 2004). Future work will entail 3 Specific Aims as follows: 1. Determine to what extent the ATP synthasome alone catalyzes partial reactions characteristic of the terminal steps of oxidative phosphorylation in mitochondria, assess the functional significance of placing PIC and ANC near the ATP synthase's motor components, and determine whether PIC and ANC form a heterodimer. 2. Elucidate at molecular resolution a topological map in 3-D in which those 8 subunit types (PIC, ANC, d, e, f, g, F6 & A6L), unique to the mitochondrial ATP synthasome, can be visualized relative to each other and to the F1 and F0 motor components common to all ATP synthases. 3. Vigorously pursue efforts in 2 different laboratories, that of the P.I. and that of J. Deisenhofer, to obtain 3-D crystals of the ATP synthasome that diffract to atomic resolution. Collectively, these novel studies are fundamental to understanding mitochondrial function in mammalian cells and have direct relevance both to identifying new effective cancer therapies and to understanding mitochondrial associated diseases. These studies also have direct relevance to using nanotechnology in medicine as the ATP synthasome is comprised of 2 reversible nanomotors.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA010951-39
Application #
7413358
Study Section
Biochemistry and Biophysics of Membranes Study Section (BBM)
Program Officer
Knowlton, John R
Project Start
1979-05-01
Project End
2010-03-31
Budget Start
2008-04-01
Budget End
2009-03-31
Support Year
39
Fiscal Year
2008
Total Cost
$388,754
Indirect Cost
Name
Johns Hopkins University
Department
Biochemistry
Type
Schools of Medicine
DUNS #
001910777
City
Baltimore
State
MD
Country
United States
Zip Code
21218
Azevedo-Silva, J; Queirós, O; Baltazar, F et al. (2016) The anticancer agent 3-bromopyruvate: a simple but powerful molecule taken from the lab to the bedside. J Bioenerg Biomembr 48:349-62
Block, Keith I; Gyllenhaal, Charlotte; Lowe, Leroy et al. (2015) Designing a broad-spectrum integrative approach for cancer prevention and treatment. Semin Cancer Biol 35 Suppl:S276-S304
Majkowska-Skrobek, Gra?yna; Augustyniak, Daria; Lis, Pawe? et al. (2014) Killing multiple myeloma cells with the small molecule 3-bromopyruvate: implications for therapy. Anticancer Drugs 25:673-82
Whitehurst, Christopher B; Sanders, Marcia K; Law, Mankit et al. (2013) Maribavir inhibits Epstein-Barr virus transcription through the EBV protein kinase. J Virol 87:5311-5
Dylag, Mariusz; Lis, Pawel; Niedzwiecka, Katarzyna et al. (2013) 3-Bromopyruvate: a novel antifungal agent against the human pathogen Cryptococcus neoformans. Biochem Biophys Res Commun 434:322-7
Darpolor, Moses M; Kaplan, David E; Pedersen, Peter L et al. (2012) Human Hepatocellular Carcinoma Metabolism: Imaging by Hyperpolarized 13C Magnetic Resonance Spectroscopy. J Liver Disease Transplant 1:
Queirós, Odília; Preto, Ana; Pacheco, António et al. (2012) Butyrate activates the monocarboxylate transporter MCT4 expression in breast cancer cells and enhances the antitumor activity of 3-bromopyruvate. J Bioenerg Biomembr 44:141-53
Ko, Y H; Verhoeven, H A; Lee, M J et al. (2012) A translational study ""case report"" on the small molecule ""energy blocker"" 3-bromopyruvate (3BP) as a potent anticancer agent: from bench side to bedside. J Bioenerg Biomembr 44:163-70
Lis, Pawe?; Zarzycki, Marek; Ko, Young H et al. (2012) Transport and cytotoxicity of the anticancer drug 3-bromopyruvate in the yeast Saccharomyces cerevisiae. J Bioenerg Biomembr 44:155-61
Blum, David J; Ko, Young H; Pedersen, Peter L (2012) Mitochondrial ATP synthase catalytic mechanism: a novel visual comparative structural approach emphasizes pivotal roles for Mg²? and P-loop residues in making ATP. Biochemistry 51:1532-46

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