title: A novel crystal form of pyrrolysyl-tRNA synthetase reveals the pre- and post-aminoacyl-tRNA synthesis conformational states of the adenylate and aminoacyl moieties and an asparagine residue in the catalytic site.
abstract: Structures of Methanosarcina mazei pyrrolysyl-tRNA synthetase (PylRS) have been determined in a novel crystal form. The triclinic form crystals contained two PylRS dimers (four monomer molecules) in the asymmetric unit, in which the two subunits in one dimer each bind N(ℇ)-(tert-butyloxycarbonyl)-L-lysyladenylate (BocLys-AMP) and the two subunits in the other dimer each bind AMP. The BocLys-AMP molecules adopt a curved conformation and the C(α) position of BocLys-AMP protrudes from the active site. The β7-β8 hairpin structures in the four PylRS molecules represent distinct conformations of different states of the aminoacyl-tRNA synthesis reaction. Tyr384, at the tip of the β7-β8 hairpin, moves from the edge to the inside of the active-site pocket and adopts multiple conformations in each state. Furthermore, a new crystal structure of the BocLys-AMPPNP-bound form is also reported. The bound BocLys adopts an unusually bent conformation, which differs from the previously reported structure. It is suggested that the present BocLys-AMPPNP-bound, BocLys-AMP-bound and AMP-bound complexes represent the initial binding of an amino acid (or pre-aminoacyl-AMP synthesis), pre-aminoacyl-tRNA synthesis and post-aminoacyl-tRNA synthesis states, respectively. The conformational changes of Asn346 that accompany the aminoacyl-tRNA synthesis reaction have been captured by X-ray crystallographic analyses. The orientation of the Asn346 side chain, which hydrogen-bonds to the carbonyl group of the amino-acid substrate, shifts by a maximum of 85-90° around the C(β) atom.
authors.affiliation: RIKEN Systems and Structural Biology Center, Tsurumi, Yokohama, Japan.
authors.forename: Tatsuo
authors.initials: T
authors.lastname: Yanagisawa
authors.forename: Tomomi
authors.initials: T
authors.lastname: Sumida
authors.forename: Ryohei
authors.initials: R
authors.lastname: Ishii
authors.forename: Shigeyuki
authors.initials: S
authors.lastname: Yokoyama
mesh_terms: D020816:Amino Acid Motifs; D000604:Amino Acyl-tRNA Synthetases / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism; D049148:Aminoacylation; D001216:Asparagine / Q000737:chemistry* / Q000378:metabolism; D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D006860:Hydrogen Bonding; D008239:Lysine / Q000031:analogs & derivatives / Q000737:chemistry; D017020:Methanosarcina / Q000201:enzymology*; D011485:Protein Binding; D011487:Protein Conformation; D055503:Protein Multimerization; D013379:Substrate Specificity
chemical_list: D001216:Asparagine; D000604:Amino Acyl-tRNA Synthetases; C456839:pyrrolysine; D008239:Lysine