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The importance of threonine‐301 from cytochromes P‐450 (laurate (ω‐1)‐hydroxylase and testosterone 16α‐hydroxylase) in substrate binding as demonstrated by site‐directed mutagenesis
Author(s) -
Imai Yoshio,
Nakamura Masahiko
Publication year - 1988
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/0014-5793(88)80106-6
Subject(s) - mutagenesis , biochemistry , chemistry , threonine , mutant , site directed mutagenesis , substrate (aquarium) , heme , histidine , binding site , enzyme , stereochemistry , biology , serine , ecology , gene
Threonine‐301 from rabbit liver cytochromes P‐450 (laurate (ω‐1)‐hydroxylase and testosterone 16α‐hydroxylase) has been replaced by histidine via site‐directed mutagenesis. In the oxidized state the mutant P‐450s exhibited typical low‐spin type absorption spectra of P‐450 and their reduced CO complexes showed a Soret peak at 450 nm. However, no spectral change was induced on addition of substrates for their wild‐type counterparts. The mutant P‐450s were also completely devoid of the hydroxylase activity. These findings suggest that threonine‐301, which is highly conserved in P‐450s and located at the distal heme surface, plays an important role in substrate binding.