2ZCE

Crystal structure of the catalytic domain of pyrrolysyl-tRNA synthetase in complex with pyrrolysine and an ATP analogue

Summary for 2ZCE

• Entry DOI: 10.2210/pdb2zce/pdb
• Related: 2E3C 2ZCD
• Descriptor: Pyrrolysyl-tRNA synthetase, MAGNESIUM ION, PYRROLYSINE, ... (5 entities in total)
• Functional Keywords: aminoacyl-trna synthetase, pyrrolysyl-trna synthetase, trna, pyrrolysine, protein3000, atp-binding, ligase, nucleotide-binding, protein biosynthesis, structural genomics, nppsfa, national project on protein structural and functional analyses, riken structural genomics/proteomics initiative, rsgi
• Biological source: Methanosarcina mazei
• Cellular location: Cytoplasm (By similarity): Q8PWY1
• Total number of polymer chains: 1
• Total formula weight: 34154.28

Authors

Yanagisawa, T.,Ishii, R.,Yokoyama, S.,RIKEN Structural Genomics/Proteomics Initiative (RSGI) (deposition date: 2007-11-08, release date: 2008-04-22, Last modification date: 2024-03-13)

Primary citation

Yanagisawa, T.,Ishii, R.,Fukunaga, R.,Kobayashi, T.,Sakamoto, K.,Yokoyama, S.
Crystallographic Studies on Multiple Conformational States of Active-site Loops in Pyrrolysyl-tRNA Synthetase
J.Mol.Biol., 378:634-652, 2008

PubMed Abstract: Pyrrolysine, a lysine derivative with a bulky pyrroline ring, is the "22nd" genetically encoded amino acid. In the present study, the carboxy-terminal catalytic fragment of Methanosarcina mazei pyrrolysyl-tRNA synthetase (PylRS) was analyzed by X-ray crystallography and site-directed mutagenesis. The catalytic fragment ligated tRNA(Pyl) with pyrrolysine nearly as efficiently as the full-length PylRS. We determined the crystal structures of the PylRS catalytic fragment in the substrate-free, ATP analogue (AMPPNP)-bound, and AMPPNP/pyrrolysine-bound forms, and compared them with the previously-reported PylRS structures. The ordering loop and the motif-2 loop undergo conformational changes from the "open" states to the "closed" states upon AMPPNP binding. On the other hand, the beta 7-beta 8 hairpin exhibits multiple conformational states, the open, intermediate (beta 7-open/beta 8-open and beta 7-closed/beta 8-open), and closed states, which are not induced upon substrate binding. The PylRS structures with a docked tRNA suggest that the active-site pocket can accommodate the CCA terminus of tRNA when the motif-2 loop is in the closed state and the beta 7-beta 8 hairpin is in the open or intermediate state. The entrance of the active-site pocket is nearly closed in the closed state of the beta 7-beta 8 hairpin, which may protect the pyrrolysyladenylate intermediate in the absence of tRNA(Pyl). Moreover, a structure-based mutational analysis revealed that hydrophobic residues in the amino acid-binding tunnel are important for accommodating the pyrrolysine side chain and that Asn346 is essential for anchoring the side-chain carbonyl and alpha-amino groups of pyrrolysine. In addition, a docking model of PylRS with tRNA was constructed based on the aspartyl-tRNA synthetase/tRNA structure, and was confirmed by a mutational analysis.

PubMed: 18387634
DOI: 10.1016/j.jmb.2008.02.045

Experimental method

X-RAY DIFFRACTION (1.8 Å)