Subtilisin secreted from a variety of Bacillus species is an alkaline protease. The peptide-chain folding of this kind of enzyme is totally different from the mammalian serine protease, but their catalytic residues ar...Subtilisin secreted from a variety of Bacillus species is an alkaline protease. The peptide-chain folding of this kind of enzyme is totally different from the mammalian serine protease, but their catalytic residues are the same and have almost identical space arrangement, so they belong to different kinds of serine proteases. The structure and function of subtilisin have been studied extensively. Subtilisin also has some other advantages for protein engineering studies and is an important industrial enzyme which is mass-used as a component of synthetic detergent to degrade proteins. Therefore a展开更多
A mutant library of subtilisin E containing random combinations of various mutagenized sites wasconstructed by one-round mutagenesis with 15 mutagenic oligonucleotides. Mutants were screened through dot blot hybridiza...A mutant library of subtilisin E containing random combinations of various mutagenized sites wasconstructed by one-round mutagenesis with 15 mutagenic oligonucleotides. Mutants were screened through dot blot hybridization and DNA sequencing. A single-point mutant (Met 222Ala) and a three-point (Asn 76Asp/Asnl09Ser/ I le 205/Cys) mutant gene from the library were expressed. The mutant proteins exhibited conspicuously improved resistance to oxidation and heat treatment, as reported before. The results show that the library is reliable and very useful for protease subtilisin E engineering.展开更多
基金Project supported by the National High-Tech. Program of China.
文摘Subtilisin secreted from a variety of Bacillus species is an alkaline protease. The peptide-chain folding of this kind of enzyme is totally different from the mammalian serine protease, but their catalytic residues are the same and have almost identical space arrangement, so they belong to different kinds of serine proteases. The structure and function of subtilisin have been studied extensively. Subtilisin also has some other advantages for protein engineering studies and is an important industrial enzyme which is mass-used as a component of synthetic detergent to degrade proteins. Therefore a
基金Project supported by United Nations Industrial Development Organization (UNIDO).
文摘A mutant library of subtilisin E containing random combinations of various mutagenized sites wasconstructed by one-round mutagenesis with 15 mutagenic oligonucleotides. Mutants were screened through dot blot hybridization and DNA sequencing. A single-point mutant (Met 222Ala) and a three-point (Asn 76Asp/Asnl09Ser/ I le 205/Cys) mutant gene from the library were expressed. The mutant proteins exhibited conspicuously improved resistance to oxidation and heat treatment, as reported before. The results show that the library is reliable and very useful for protease subtilisin E engineering.
