Active Glutaminase C Self-assembles into a Supratetrameric Oligomer That Can Be Disrupted by an Allosteric Inhibitor

Author(s):Show less -

Scota Ferreira, Amanda Petrina ^[1] ; Cassago, Alexandre ^[2] ; Goncalves, Kaliandra de Almeida ^[1] ; Dias, Marilia Meira ^[1] ; Adamoski, Douglas ^[1] ; Rodrigues Ascencao, Carolline Fernanda ^[1] ; Honorato, Rodrigo Vargas ^[1] ; de Oliveira, Juliana Ferreira ^[1] ; Ferreira, Igor Monteze ^[1] ; Fornezari, Camila ^[1] ; Bettini, Jefferson ^[2] ; Lopes Oliveira, Paulo Sergio ^[1] ; Paes Leme, Adriana Franco ^[1] ; Portugal, Rodrigo Villares ^[2] ; Berteli Ambrosio, Andre Luis ^[1] ; Gomes Dias, Sandra Martha ^[1]

Affiliation

^[1] CNPEM, Lab Nacionais Biociencias, BR-13083100 Campinas, SP - Brazil

^[2] CNPEM, BR-13083100 Campinas, SP - Brazil

• Document type: Journal article
• Source: Journal of Biological Chemistry; v. 288, n. 39, p. 28009-28020, SEP 27 2013.
• Web of Science Citations: 28

Abstract

The phosphate-dependent transition between enzymatically inert dimers into catalytically capable tetramers has long been the accepted mechanism for the glutaminase activation. Here, we demonstrate that activated glutaminase C(GAC) self-assembles into a helical, fiber-like double-stranded oligomer and propose a molecular model consisting of seven tetramer copies per turn per strand interacting via the N-terminal domains. The loop (LRFNKL326)-L-321 is projected as the major regulating element for self-assembly and enzyme activation. Furthermore, the previously identified in vivo lysine acetylation (Lys(311) in humans, Lys(316) in mouse) is here proposed as an important down-regulator of superoligomer assembly and protein activation. Bis-2-(5-phenylacetamido-1,3,4-thiadiazol-2-yl) ethyl sulfide, a known glutaminase inhibitor, completely disrupted the higher order oligomer, explaining its allosteric mechanism of inhibition via tetramer stabilization. A direct correlation between the tendency to self-assemble and the activity levels of the three mammalian glutaminase isozymes was established, with GAC being the most active enzyme while forming the longest structures. Lastly, the ectopic expression of a fiber-prone superactive GAC mutant in MDA-MB 231 cancer cells provided considerable proliferative advantages to transformed cells. These findings yield unique implications for the development of GAC-oriented therapeutics targeting tumor metabolism. (AU)

• FAPESP's process: 12/14298-9 - Cancer metabolic adaptation: structural and functional studies of key proteins
• Grantee: Andre Luis Berteli Ambrosio
• Support Opportunities: Regular Research Grants
• FAPESP's process: 10/05003-0 - Structural and functional studies of key proteins underlying the metabolic adaptation process in tumors
• Grantee: Andre Luis Berteli Ambrosio
• Support Opportunities: Regular Research Grants
• FAPESP's process: 09/10875-9 - Cellular and biochemical studies of the glutaminase enzyme and its relation with cancer
• Grantee: Sandra Martha Gomes Dias
• Support Opportunities: Research Grants - Young Investigators Grants
• FAPESP's process: 10/05987-0 - Structural determination of the kidney type glutaminase and the search for its binding partners.
• Grantee: Alexandre Cassago
• Support Opportunities: Scholarships in Brazil - Post-Doctoral
• FAPESP's process: 09/54067-3 - Acquisition of a mass spectrometer coupled to a liquid chromatography system for increasing the capacity to meet the needs of users and for making new technologies available in the Laboratory of Mass Spectrometry
• Grantee: Adriana Franco Paes Leme
• Support Opportunities: Multi-user Equipment Program
• FAPESP's process: 11/06654-7 - Screening of natural extracts for glycolytic and glutaminolytic inhibitory properties as a platform for searching new anti-tumours compounds
• Grantee: Kaliandra de Almeida Gonçalves
• Support Opportunities: Scholarships in Brazil - Post-Doctoral