title: Computational protein design with a generalized Born solvent model: application to Asparaginyl-tRNA synthetase.
abstract: Computational Protein Design (CPD) is a promising method for high throughput protein and ligand mutagenesis. Recently, we developed a CPD method that used a polar-hydrogen energy function for protein interactions and a Coulomb/Accessible Surface Area (CASA) model for solvent effects. We applied this method to engineer aspartyl-adenylate (AspAMP) specificity into Asparaginyl-tRNA synthetase (AsnRS), whose substrate is asparaginyl-adenylate (AsnAMP). Here, we implement a more accurate function, with an all-atom energy for protein interactions and a residue-pairwise generalized Born model for solvent effects. As a first test, we compute aminoacid affinities for several point mutants of Aspartyl-tRNA synthetase (AspRS) and Tyrosyl-tRNA synthetase and stability changes for three helical peptides and compare with experiment. As a second test, we readdress the problem of AsnRS aminoacid engineering. We compare three design criteria, which optimize the folding free-energy, the absolute AspAMP affinity, and the relative (AspAMP-AsnAMP) affinity. The sequences and conformations are improved with respect to our previous, polar-hydrogen/CASA study: For several designed complexes, the AspAMP carboxylate forms three interactions with a conserved arginine and a designed lysine, as in the active site of the AspRS:AspAMP complex. The conformations and interactions are well maintained in molecular dynamics simulations and the sequences have an inverted specificity, favoring AspAMP over AsnAMP. The method is not fully successful, since experimental measurements with the seven most promising sequences show that they do not catalyze at a detectable level the adenylation of Asp (or Asn) with ATP. This may be due to weak AspAMP binding and/or disruption of transition-state stabilization.
authors.affiliation: Department of Physics, University of Cyprus, Nicosia, Cyprus.
authors.forename: Savvas
authors.initials: S
authors.lastname: Polydorides
authors.forename: Najette
authors.initials: N
authors.lastname: Amara
authors.forename: Caroline
authors.initials: C
authors.lastname: Aubard
authors.forename: Pierre
authors.initials: P
authors.lastname: Plateau
authors.forename: Thomas
authors.initials: T
authors.lastname: Simonson
authors.forename: Georgios
authors.initials: G
authors.lastname: Archontis
mesh_terms: D000595:Amino Acid Sequence; D000596:Amino Acids / Q000737:chemistry / Q000378:metabolism; D001226:Aspartate-tRNA Ligase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites; D019295:Computational Biology / Q000379:methods*; D008024:Ligands; D008958:Models, Molecular; D056004:Molecular Dynamics Simulation; D017354:Point Mutation; D011485:Protein Binding; D011487:Protein Conformation; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D012346:RNA, Transfer, Amino Acyl / Q000737:chemistry* / Q000378:metabolism*; D013379:Substrate Specificity; D014448:Tyrosine-tRNA Ligase / Q000737:chemistry / Q000235:genetics
chemical_list: D000596:Amino Acids; D008024:Ligands; D012346:RNA, Transfer, Amino Acyl; D014448:Tyrosine-tRNA Ligase; D001226:Aspartate-tRNA Ligase; C029194:asparaginyl-tRNA synthetase