Bacteria readily oxidize alkanes utilizing the metalloenzyme alkane hydroxylase (AlkB) and thus the actions ofAlkB represent one of nature?s best defenses against the environmental effects of oil spills. Mammalian fattyacids are also desaturated by integral membrane fatty acid desaturases that are structurally quite similar to AlkB.It is not understood why AlkB catalyzes the addition of an OH group to an alkane while desaturases convertsingle C-C bonds to double C=C bonds. AlkB and soluble methane monooxygenase (sMMO) both catalyze thetransformation of inert C-H bonds using a diiron catalyst but AlkB has a diiron catalyst with primarily nitrogenatoms as ligands while sMMO has an oxygen-rich coordination site. The coordination environment is predictedto influence the electronic structure and hence reactivity but the electronic structure of AlkB has not beencharacterized. The long-term objective of this research effort is to understand how the structure of AlkBdetermines the chemistry it affects and to use this knowledge to deepen our understanding of how biologyselectively activates molecular oxygen and catalyzes the oxidation of inert hydrocarbons. An associatedobjective is to understand how structurally very similar biological motifs select between catalyzing thehydroxylation of alkanes or their desaturation. The primary goal of the specific research proposed is obtain awell-diffracting crystal of AlkB that will enable us to determine the three-dimensional structure of this enzyme.Related goals are to spectroscopically characterize the ground state and reactive intermediates and to beginexplore AlkBs and related metalloenzymes from additional microorganisms. These goals enable us to test thehypothesis that the structures of the active sites of all AlkBs and membrane-spanning desaturases are the samebut that structural differences in the substrate binding pocket control substrate selectivity and catalyst reactivity.If successful, this work would have an impact on bioinorganic chemistry and environmental chemistry. A three-dimensional structure of AlkB would help to answer many questions about the range of chemical motifs thatbiology can use to oxidize alkanes. It would also answer questions about some of the chemical processesutilized in the environmental response to oil spills.
Principal Investigator: Austin; Rachel NarehoodProject NarrativeThis project focuses on understanding how the enzyme that catalyzes the first step in thetransformation of oil to carbon dioxide works. It will also explore how this enzyme; which isstructurally similar to enzymes important in human lipid metabolism; functions in comparisonwith that important class of enzymes as well.
