title: Engineering photocycle dynamics. Crystal structures and kinetics of three photoactive yellow protein hinge-bending mutants.
abstract: Crystallographic and spectroscopic analyses of three hinge-bending mutants of the photoactive yellow protein are described. Previous studies have identified Gly(47) and Gly(51) as possible hinge points in the structure of the protein, allowing backbone segments around the chromophore to undergo large concerted motions. We have designed, crystallized, and solved the structures of three mutants: G47S, G51S, and G47S/G51S. The protein dynamics of these mutants are significantly affected. Transitions in the photocycle, measured with laser induced transient absorption spectroscopy, show rates up to 6-fold different from the wild type protein and show an additive effect in the double mutant. Compared with the native structure, no significant conformational differences were observed in the structures of the mutant proteins. We conclude that the structural and dynamic integrity of the region around these mutations is of crucial importance to the photocycle and suggest that the hinge-bending properties of Gly(51) may also play a role in PAS domain proteins where it is one of the few conserved residues.
authors.affiliation: W. M. Keck Structural Biology, Cold Spring Harbor Laboratory, Cold Spring Harbor, New York 11724, USA. dava@davapc1.bioch.dundee.ac.uk
authors.forename: Daan M F
authors.initials: DM
authors.lastname: van Aalten
authors.forename: Andrea
authors.initials: A
authors.lastname: Haker
authors.forename: Johnny
authors.initials: J
authors.lastname: Hendriks
authors.forename: Klaas J
authors.initials: KJ
authors.lastname: Hellingwerf
authors.forename: Leemor
authors.initials: L
authors.lastname: Joshua-Tor
authors.forename: Wim
authors.initials: W
authors.lastname: Crielaard
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D018360:Crystallography, X-Ray; D004926:Escherichia coli; D020563:Gammaproteobacteria / Q000378:metabolism / Q000528:radiation effects; D007700:Kinetics; D008027:Light; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D020283:Photoreceptors, Microbial; D011487:Protein Conformation
chemical_list: D001426:Bacterial Proteins; D020283:Photoreceptors, Microbial; C050162:photoactive yellow protein, Bacteria