title: Structure, function and inhibition of the two- and three-domain 4Fe-4S IspG proteins.
abstract: IspG is a 4Fe4S protein involved in isoprenoid biosynthesis. Most bacterial IspGs contain two domains: a TIM barrel (A) and a 4Fe4S domain (B), but in plants and malaria parasites, there is a large insert domain (A*) whose structure and function are unknown. We show that bacterial IspGs function in solution as (AB)(2) dimers and that mutations in either both A or both B domains block activity. Chimeras harboring an A-mutation in one chain and a B-mutation in the other have 50% of the activity seen in wild-type protein, because there is still one catalytically active AB domain. However, a plant IspG functions as an AA*B monomer. We propose, using computational modeling and electron microscopy, that the A* insert domain has a TIM barrel structure that interacts with the A domain. This structural arrangement enables the A and B domains to interact in a "cup and ball" manner during catalysis, just as in the bacterial systems. EPR/HYSCORE spectra of reaction intermediate, product, and inhibitor ligands bound to both two and three domain proteins are identical, indicating the same local electronic structure, and computational docking indicates these ligands bridge both A and B domains. Overall, the results are of broad general interest because they indicate the insert domain in three-domain IspGs is a second TIM barrel that plays a structural role and that the pattern of inhibition of both two and three domain proteins are the same, results that can be expected to be of use in drug design.
authors.affiliation: Center for Biophysics and Computational Biology, University of Illinois, Urbana, IL 61801, USA.
authors.forename: Yi-Liang
authors.initials: YL
authors.lastname: Liu
authors.forename: Francisco
authors.initials: F
authors.lastname: Guerra
authors.forename: Ke
authors.initials: K
authors.lastname: Wang
authors.forename: Weixue
authors.initials: W
authors.lastname: Wang
authors.forename: Jikun
authors.initials: J
authors.lastname: Li
authors.forename: Cancan
authors.initials: C
authors.lastname: Huang
authors.forename: Wei
authors.initials: W
authors.lastname: Zhu
authors.forename: Kevin
authors.initials: K
authors.lastname: Houlihan
authors.forename: Zhi
authors.initials: Z
authors.lastname: Li
authors.forename: Yong
authors.initials: Y
authors.lastname: Zhang
authors.forename: Satish K
authors.initials: SK
authors.lastname: Nair
authors.forename: Eric
authors.initials: E
authors.lastname: Oldfield
mesh_terms: D000480:Alkynes / Q000737:chemistry / Q000494:pharmacology; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D001665:Binding Sites / Q000235:genetics; D055162:Biocatalysis / Q000187:drug effects; D003198:Computer Simulation; D011756:Diphosphates / Q000737:chemistry / Q000494:pharmacology; D004578:Electron Spin Resonance Spectroscopy; D007506:Iron-Sulfur Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008024:Ligands; D008854:Microscopy, Electron; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D011485:Protein Binding / Q000187:drug effects; D055503:Protein Multimerization; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid
chemical_list: D000480:Alkynes; D001426:Bacterial Proteins; D011756:Diphosphates; D007506:Iron-Sulfur Proteins; D008024:Ligands