title: Small-angle X-ray scattering study of a Rex family repressor: conformational response to NADH and NAD+ binding in solution.
abstract: The transcriptional repressor Rex is a sensor of the intracellular NADH/NAD(+) redox state through direct binding of NADH or NAD(+). Homodimeric Rex protein from Thermus aquaticus (T-Rex) and Bacillus subtilis (B-Rex) exists in several different conformations. In both organisms, Rex in complex with NADH has the DNA binding domains packed together at the dimer interface, whereas in the apo form of B-Rex the linkers connecting these domains to the core are flexible. The crystal structures of the apo forms of B-Rex and a mutated variant of T-Rex are radically different. We describe the solution structures of B-Rex in complex with NAD(+) or NADH and in its apo form, on the basis of small-angle X-ray scattering (SAXS) measurements. This study addresses to what extent the unusual orientation of the DNA recognition domains of the crystal structure of apo B-Rex is due to stabilization by crystal packing. Low-resolution ab initio solution structures were obtained for apo B-Rex, B-Rex:NADH and B-Rex:NAD(+). Models giving a more detailed picture of these three solution structures were obtained also by rigid body fitting of the crystallographic domains. The SAXS data confirm the elongated and flexible nature of apo-B-Rex and the existence of two distinct and more rigid conformations for the complexes with NADH and NAD(+). The models emerging from this study indicate a reaction mechanism for B-Rex in which the recognition domains are rotated upon binding to NADH.
authors.affiliation: Department of Biochemistry and Structural Biology, Lund University, SE-22100 Lund, Sweden.
authors.forename: Ellen
authors.initials: E
authors.lastname: Wang
authors.forename: Teemu P
authors.initials: TP
authors.lastname: Ikonen
authors.forename: Matti
authors.initials: M
authors.lastname: Knaapila
authors.forename: Dmitri
authors.initials: D
authors.lastname: Svergun
authors.forename: Derek T
authors.initials: DT
authors.lastname: Logan
authors.forename: Claes
authors.initials: C
authors.lastname: von Wachenfeldt
mesh_terms: D000595:Amino Acid Sequence; D001412:Bacillus subtilis / Q000378:metabolism; D001426:Bacterial Proteins / Q000737:chemistry*; D001665:Binding Sites; D018360:Crystallography, X-Ray; D009243:NAD / Q000737:chemistry; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry; D012097:Repressor Proteins / Q000737:chemistry*; D053838:Scattering, Small Angle; D012996:Solutions / Q000737:chemistry; D013824:Thermus / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D011994:Recombinant Proteins; D012097:Repressor Proteins; D012996:Solutions; D009243:NAD