Computational protocol: UPF201 Archaeal Specific Family Members Reveal Structural Similarity to RNA-Binding Proteins but Low Likelihood for RNA-Binding Function

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Protocol publication

[…] All structures were determined independently via single wavelength anomalous dispersion (SAD) with Se-Met crystals. In each case, SAD data collection at an X-ray wavelength corresponding to the crystal Se emission line sufficed for determining the Se atom substructure with SHELXD . For 10077a, crystals were obtained in a triclinic space group with 4 molecules in the asymmetric unit, and the structure could only be determined after combining two full-sphere SAD data sets recorded from two crystals. Initial phases were obtained with SHARP , and further improved via density modification using DM . In all cases, about 70% of the polypeptide chain was built automatically by ARP/wARP except in the case of 10077d where the data extended to 3 Å only. Subsequent model building was performed manually using O . Structure refinement was performed with simulated annealing followed by Powell energy minimization . The refined atomic model was evaluated using the RCSB AUTODEP deposit tool ( Final refinement statistics are given in . [...] 1) Secondary structural elements, hydrogen bonds, solvent accessible surface area, buried residues, and folding free energy were calculated using VADAR . 2) Ionic interactions (salt bridges) and cation-pi interactions were calculated using PIC . 3) Secondary structure attribution of residues and hydrogen bonds were calculated using DSSP . 4) Contact and potential energies were calculated with PSQS . 5) To calculate geometry of the probable protease, glycolysis pathway enzyme, or metal binding motif residues PAR-3D ( was used, though none was identified . 6) Putative RNA binding residues were identified using BindN (, RNAbindR (, and KYG ( –. 7) Conserved residues were mapped onto the structure using ConSurf . 8) For phylogenetic analysis the structure based multiple sequence alignment and the resulting tree was constructed using 3Dcoffee choice from the T-coffee package ( . […]

Pipeline specifications

Software tools SHELX, ARP/wARP, VADAR, BindN, RNABindR, KYG, ConSurf, T-Coffee
Applications Phylogenetics, Protein structure analysis
Organisms Dipturus trachyderma