This review summarizes the subtilisin-like serine protease superfamily, which includes over 200 known members, with more than 100 newly identified in the past five years. These enzymes are found in Archaea, Bacteria, fungi, yeasts, and higher eukaryotes. Subtilases are characterized by their catalytic domains, which are highly conserved and contain residues such as Asp32, His64, and Ser221. The catalytic domain is typically located at the N-terminal end of the enzyme, following the pre-pro region. Subtilases are synthesized as pre-pro-enzymes, translocated across cell membranes, and activated by cleavage of the pro-peptide. The subtilase superfamily is divided into six families based on sequence homology. These families include the subtilisin family, thermitase family, proteinase K family, lantibiotic peptidase family, kexin family, and pyrolysin family. Each family has distinct characteristics, such as sequence identity, structural features, and substrate specificity. The subtilisin family is mainly found in microorganisms and includes enzymes from Bacillus, with subgroups of true subtilisins, high-alkaline proteases, and intracellular proteases. The thermitase family is also found in microorganisms, including some thermophiles and halophiles. The proteinase K family is found in fungi, yeasts, and gram-negative bacteria, with a high degree of sequence similarity. The lantibiotic peptidase family is found in gram-positive bacteria and is characterized by low sequence similarity with other subtilases. The kexin family is involved in the activation of peptide hormones, growth factors, and viral proteins. The pyrolysin family is a heterogeneous group of enzymes with low sequence conservation. The catalytic domains of subtilases are highly conserved, with several residues being essential for catalytic activity. These residues include the catalytic triad (Asp32, His64, and Ser221) and the oxyanion-hole residue (Asn155). The catalytic domain is typically surrounded by variable regions, which are involved in substrate binding and enzyme-substrate interactions. The subtilase superfamily is also characterized by large insertions and deletions, which can affect the enzyme's structure and function. These insertions and deletions are often found in the variable regions and can influence the enzyme's substrate specificity and catalytic efficiency. The subtilase superfamily is also characterized by the presence of calcium-binding sites, which are essential for the enzyme's stability and activity. These calcium-binding sites are typically located in the catalytic domain and are involved in the enzyme's function. The subtilase superfamily is also characterized by the presence of disulfide bonds, which contribute to the enzyme's overall stability.This review summarizes the subtilisin-like serine protease superfamily, which includes over 200 known members, with more than 100 newly identified in the past five years. These enzymes are found in Archaea, Bacteria, fungi, yeasts, and higher eukaryotes. Subtilases are characterized by their catalytic domains, which are highly conserved and contain residues such as Asp32, His64, and Ser221. The catalytic domain is typically located at the N-terminal end of the enzyme, following the pre-pro region. Subtilases are synthesized as pre-pro-enzymes, translocated across cell membranes, and activated by cleavage of the pro-peptide. The subtilase superfamily is divided into six families based on sequence homology. These families include the subtilisin family, thermitase family, proteinase K family, lantibiotic peptidase family, kexin family, and pyrolysin family. Each family has distinct characteristics, such as sequence identity, structural features, and substrate specificity. The subtilisin family is mainly found in microorganisms and includes enzymes from Bacillus, with subgroups of true subtilisins, high-alkaline proteases, and intracellular proteases. The thermitase family is also found in microorganisms, including some thermophiles and halophiles. The proteinase K family is found in fungi, yeasts, and gram-negative bacteria, with a high degree of sequence similarity. The lantibiotic peptidase family is found in gram-positive bacteria and is characterized by low sequence similarity with other subtilases. The kexin family is involved in the activation of peptide hormones, growth factors, and viral proteins. The pyrolysin family is a heterogeneous group of enzymes with low sequence conservation. The catalytic domains of subtilases are highly conserved, with several residues being essential for catalytic activity. These residues include the catalytic triad (Asp32, His64, and Ser221) and the oxyanion-hole residue (Asn155). The catalytic domain is typically surrounded by variable regions, which are involved in substrate binding and enzyme-substrate interactions. The subtilase superfamily is also characterized by large insertions and deletions, which can affect the enzyme's structure and function. These insertions and deletions are often found in the variable regions and can influence the enzyme's substrate specificity and catalytic efficiency. The subtilase superfamily is also characterized by the presence of calcium-binding sites, which are essential for the enzyme's stability and activity. These calcium-binding sites are typically located in the catalytic domain and are involved in the enzyme's function. The subtilase superfamily is also characterized by the presence of disulfide bonds, which contribute to the enzyme's overall stability.