Monday, June 18, 2007 Golden Eagle Eyrie (Boise Centre on the Grove) 144
Mechanistic analysis of an Archaeal antioxidant protein cage by mass spectrometry
Luke M. Oltrogge and Trevor Douglas. Montana State University, Bozeman, MT
Oxidative stress poses distinct problems that aerobic organisms constantly face. Sulfolobus solfataricus, a hyperthermo-acidophilic archaeon, combats reactive oxygen species (ROS) using a protein cage from a recently discovered class of enzymes, Dps-L proteins. The active site of SsDps-L located deep within a 4-α-helix bundle and contains a dimetal binding site. Approximately 8Å above the active site lies a cysteine pair that is highly conserved among Dps-L proteins. It is thought that these cysteines may play an active role in the reduction-oxidation cycle or function in a regulatory capacity. In addition, the activity of SsDps-L is dependent upon the metals ions bound in the active site and displays ferritin-like mineralization in the presence of Fe(II) and catalase-like disproportionation of peroxide in the presence of Mn2+. This study investigates the reduction-oxidation status of the cysteine pair in in vitro and in vivo systems with proteolysis and highly sensitive and specific mass spectrometry techniques. The roles of Fe(II) and Mn(II) and their binding affinities in the enzyme is also being explored through the use of non-covalent mass spectrometry in order to resolve the dual mineralization / disproportionation function of this enzyme.