title: Characterization of a Radical Intermediate in Lipoyl Cofactor Biosynthesis.
abstract: Lipoyl synthase (LipA) catalyzes the final step in the biosynthesis of the lipoyl cofactor, the insertion of two sulfur atoms at C6 and C8 of an n-octanoyl chain. LipA is a member of the radical S-adenosylmethionine (SAM) superfamily of enzymes and uses two [4Fe-4S] clusters to catalyze its transformation. One cluster binds in contact with SAM and donates the requisite electron for the reductive cleavage of SAM to generate two 5'-deoxyadenosyl 5'-radicals, which abstract hydrogen atoms from C6 and C8 of the substrate. By contrast, the second, auxiliary [4Fe-4S] cluster, has been hypothesized to serve as the sulfur donor in the reaction. Such a sacrificial role for an iron-sulfur cluster during catalysis has not been universally accepted. Use of a conjugated 2,4-hexadienoyl-containing substrate analogue has allowed the substrate radical to be trapped and characterized by continuous-wave and pulsed electron paramagnetic resonance methods. Here we report the observation of a (57)Fe hyperfine coupling interaction with the paramagnetic signal, which indicates that the iron-sulfur cluster of LipA and its substrate are within bonding distance.
authors.affiliation: Departments of †Chemistry and ‡Biochemistry and Molecular Biology and §The Huck Institutes of the Life Sciences, The Pennsylvania State University , University Park, Pennsylvania 16802, United States.
authors.forename: Nicholas D
authors.initials: ND
authors.lastname: Lanz
authors.affiliation: Departments of †Chemistry and ‡Biochemistry and Molecular Biology and §The Huck Institutes of the Life Sciences, The Pennsylvania State University , University Park, Pennsylvania 16802, United States.
authors.forename: Justin M
authors.initials: JM
authors.lastname: Rectenwald
authors.affiliation: Departments of †Chemistry and ‡Biochemistry and Molecular Biology and §The Huck Institutes of the Life Sciences, The Pennsylvania State University , University Park, Pennsylvania 16802, United States.
authors.forename: Bo
authors.initials: B
authors.lastname: Wang
authors.affiliation: Departments of †Chemistry and ‡Biochemistry and Molecular Biology and §The Huck Institutes of the Life Sciences, The Pennsylvania State University , University Park, Pennsylvania 16802, United States.
authors.forename: Elizabeth S
authors.initials: ES
authors.lastname: Kakar
authors.affiliation: Departments of †Chemistry and ‡Biochemistry and Molecular Biology and §The Huck Institutes of the Life Sciences, The Pennsylvania State University , University Park, Pennsylvania 16802, United States.
authors.forename: Tatiana N
authors.initials: TN
authors.lastname: Laremore
authors.affiliation: Departments of †Chemistry and ‡Biochemistry and Molecular Biology and §The Huck Institutes of the Life Sciences, The Pennsylvania State University , University Park, Pennsylvania 16802, United States.
authors.forename: Squire J
authors.initials: SJ
authors.lastname: Booker
authors.affiliation: Departments of †Chemistry and ‡Biochemistry and Molecular Biology and §The Huck Institutes of the Life Sciences, The Pennsylvania State University , University Park, Pennsylvania 16802, United States.
authors.forename: Alexey
authors.initials: A
authors.lastname: Silakov
mesh_terms: D004798:Enzymes / Q000737:chemistry*; D005609:Free Radicals; D013058:Mass Spectrometry; D012436:S-Adenosylmethionine / Q000737:chemistry*
chemical_list: D004798:Enzymes; D005609:Free Radicals; D012436:S-Adenosylmethionine