Computational protocol: In Silico Prediction and In Vitro Characterization of Multifunctional Human RNase3

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Protocol publication

[…] Unique peptides of query proteins, 13 hRNaseA family members, were identified employing Reinforced Merging for Unique Segments ReMUS system (ReMUS) (http://140.121.196.30/remus.asp) []. The system adopted a bottom-up strategy to extract unique patterns in each sequence at different unique levels. A fundamental unique peptide segment with previously defined pattern length, named as primary pattern was extracted at the first step. The rule of thumb for primary pattern lengths is that a shorter length setting for similar protein sequences and a longer length for dissimilar ones. The length of primary pattern in this study is set as 3 residues for hRNaseA protein family. After that Boyer Moore algorithm was performed to efficiently retrieve all primary patterns among all sequences. Each verified fundamental unique peptide segment was analyzed based on its frequencies of appearance, and its representation level of uniqueness was calculated for the merging processes in the next module. The last merging algorithm concatenated these extracted unique peptide segments through a bottom-up approach only if the primary unique peptide segments were overlapped within a sequence. The merged segments were guaranteed with unique features compared to all other protein sequences in the query dataset.Clustal W2 (http://www.ebi.ac.uk/Tools/msa/clustalw2/) was used to align protein sequences of 13 hRNaseA family members on the basis of automatically progressive alignment mode. Protein sequences were retrieved from UniProtKB (http://www.uniprot.org/). To perform multiple-sequence alignment, gap open and extend penalties were set to 10 and 0.2, respectively. For secondary structure analysis in corresponding HBRs of hRNase1 to hRNase8, tertiary structures of hRNase1, 2, 3, 4, 5, and 7 were collected from protein data bank (PDB, http://www.rcsb.org/pdb/home/home.do), and those of hRNase6 and hRNase8 were simulated by Protein Structure Prediction Server (PS)2 (http://ps2.life.nctu.edu.tw/) using hRNase7 as a template. In addition, National Center for Biotechnology Information (NCBI) Blast (http://blast.ncbi.nlm.nih.gov/Blast.cgi) was employed to compare sequence correspondent to HBPRNase3(32–41) among nonhuman primate hRNase2s and hRNase3s, as well as human RNaseA superfamily members. […]

Pipeline specifications

Software tools ReMUS, Clustal W, BLASTN
Application Protein structure analysis
Organisms Homo sapiens
Diseases Mucopolysaccharidosis III