Iron-Sulfur Clusters in Biological Radical Generation

Information

  • Research Project
  • 7692527
  • ApplicationId
    7692527
  • Core Project Number
    R01GM054608
  • Full Project Number
    3R01GM054608-12S1
  • Serial Number
    54608
  • FOA Number
  • Sub Project Id
  • Project Start Date
    8/1/1997 - 26 years ago
  • Project End Date
    5/31/2010 - 14 years ago
  • Program Officer Name
    IKEDA, RICHARD A.
  • Budget Start Date
    6/1/2008 - 16 years ago
  • Budget End Date
    5/31/2009 - 15 years ago
  • Fiscal Year
    2008
  • Support Year
    12
  • Suffix
    S1
  • Award Notice Date
    9/29/2008 - 15 years ago
Organizations

Iron-Sulfur Clusters in Biological Radical Generation

[unreadable] DESCRIPTION (provided by applicant): The overall objective of this project is to delineate the detailed chemical mechanism of radical generation by the Fe/S-S-adenosylmethionine (the so-called radical SAM) superfamily of enzymes. These enzymes span a remarkably diverse range of reactions and appear to be represented across the phylogenetic kingdom, with hundreds of radical SAM enzymes identified. The widespread occurrence of these enzymes throughout biology, from bacteria to humans, is indicative of the significance of the chemistry catalyzed by these enzymes. In humans, radical SAM enzymes are involved in the biosynthesis of lipoic acid, the synthesis of heme, and the biosynthesis of the molybdopterin cofactor, among many other essential functions, some as yet unidentified. Despite the diversity of reactions catalyzed, our overriding hypothesis is that the adenosylmethionine-dependent iron-sulfur enzymes all operate by a common mechanism in which a reduced cluster interacts with S-adenosylmethionine to generate an adenosyl radical intermediate, which is directly involved in catalysis. These reactions represent novel chemistry for iron-sulfur clusters. To investigate this novel chemistry, biochemical, spectroscopic, mechanistic, and structural studies of pyruvate formate-lyase activating enzyme (PFL-AE) will be pursued. The specific aims include further investigation of the [4Fe-4S] cluster of PFL-AE, its unique iron site, its interactions with substrate, and its role in catalysis. In addition we propose to explore the nature of the PFL-AE iron-sulfur cluster in whole cells. We will also utilize substrate analogs as probes of ES interactions and the PFL-AE catalytic mechanism. We will use spectroscopic approaches, particularly ENDOR, to probe the structural and electronic features of the PFL- AE/AdoMet/PFL complex. We will also utilize cryoreduction and rapid-freeze-quench spectroscopic approaches to identify and spectroscopically characterize intermediates in the radical-generation reaction catalyzed by PFL-AE. Finally, we will pursue structural characterization of PFL-AE and its complexes with PFL and SAM using X-ray crystallography. [unreadable] [unreadable] [unreadable]

IC Name
NATIONAL INSTITUTE OF GENERAL MEDICAL SCIENCES
  • Activity
    R01
  • Administering IC
    GM
  • Application Type
    3
  • Direct Cost Amount
  • Indirect Cost Amount
  • Total Cost
    71250
  • Sub Project Total Cost
  • ARRA Funded
  • CFDA Code
    859
  • Ed Inst. Type
    SCHOOLS OF ARTS AND SCIENCES
  • Funding ICs
    NIGMS:71250\
  • Funding Mechanism
  • Study Section
    ZRG1
  • Study Section Name
    Special Emphasis Panel
  • Organization Name
    MONTANA STATE UNIVERSITY (BOZEMAN)
  • Organization Department
    NONE
  • Organization DUNS
  • Organization City
    BOZEMAN
  • Organization State
    MT
  • Organization Country
    UNITED STATES
  • Organization Zip Code
    59717
  • Organization District
    UNITED STATES