Computational protocol: Clustering of disulfide-rich peptides provides scaffolds for hit discovery by phage display: application to interleukin-23

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Protocol publication

[…] Native overlap was defined as the fraction of Cα atoms in one DRP that were within 3.5 Å of the corresponding atoms in a second DRP following structural alignment of the first DRP to the second DRP. Thus, a native overlap of 1.0 meant that all equivalent residues across the aligned DRPs are within 3.5 Å of each other and there are no gaps in the alignment (i.e. every residue in one DRP had an equivalent in the other). Structural alignments were performed using the iterative_structure_align() command in MODELLER version 9.10 []; this command implemented the SALIGN algorithm []. [...] For each cluster, a multiple structure alignment was performed for all DRPs using SALIGN. A multiple sequence alignment was produced based on the structure alignment and used as input to the program AL2CO [], which quantified the overall degree of conservation at each position in an alignment. The ‘sum of pairs’ method of AL2CO was used, using the BLOSUM62 scoring matrix [] to compare similar amino acid residue types. AL2CO calculated normalized scores at each position ranging from −2 to 2; these scores were scaled to RGB color values that could be used by the structure visualization program PyMol [] to color individual residues; thus, each residue was colored on a RGB scale of blue [0, 0, 255] to yellow [255, 255, 0]. Commands to perform the coloring were automatically generated and saved in a PyMol script, which read the aligned structures generated by SALIGN and colored each residue for each DRP according to the degree of sequence conservation in the alignment. […]

Pipeline specifications

Software tools MODELLER, SALIGN, AL2CO, PyMOL
Application Protein structure analysis