PSIBLAST

PSIBLAST, short for Position-Specific Iterated BLAST, is a protein sequence search tool in the BLAST family designed to detect distant protein homology. It enhances sensitivity by building and refining a position-specific scoring matrix (PSSM) from sequences aligned in an initial search and then using that matrix to search the database in subsequent iterations.

How it works. An initial BLASTP search is performed with a query protein sequence against a protein database. Sequences that meet a chosen significance threshold are aligned with the query to construct a PSSM, which captures position-specific residue substitution preferences. This PSSM is then used to perform a new search, identifying additional matches and providing updated alignments. The process repeats for a set number of iterations or until no new significant sequences are found. Default iterations are commonly around five, and users can adjust parameters such as the E-value threshold and inclusion of low-complexity filtering.

Output and interpretation. PSI-BLAST reports significant hits, E-values, and alignment details for each iteration. The resulting PSSM can be saved for further analysis, and the tool is often used to infer function, identify conserved domains, and build evolutionary profiles. It is capable of detecting remote homology that ordinary BLAST searches may miss.

Limitations. PSI-BLAST can propagate errors if early results include non-homologous sequences or biased data, and its performance depends on iteration settings and database composition. Proper interpretation and validation are advised, especially in exploratory analyses.