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Functional expression of thiocyanate hydrolase is promoted by its activator protein, P15K
Author(s) -
Kataoka Shingo,
Arakawa Takatoshi,
Hori Shota,
Katayama Yoko,
Hara Yoshiko,
Matsushita Yasuhiko,
Nakayama Hiroshi,
Yohda Masafumi,
Nyunoya Hiroshi,
Dohmae Naoshi,
Maeda Mizuo,
Odaka Masafumi
Publication year - 2006
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/j.febslet.2006.07.051
Subject(s) - cobalt , chemistry , escherichia coli , trimer , hydrolase , thiocyanate , enzyme , activator (genetics) , dodecameric protein , ligand (biochemistry) , biochemistry , stereochemistry , dimer , gene , inorganic chemistry , receptor , organic chemistry , dna
Thiocyanate hydrolase (SCNase) is a cobalt‐containing enzyme with a post‐translationally modified cysteine ligand, γCys131‐SO 2 H. When the SCNase α, β and γ subunits were expressed in Escherichia coli , the subunits assembled to form a hetero‐dodecamer, (αβγ) 4 , like native SCNase but exhibited no catalytic activity. Metal analysis indicated that SCNase was expressed as an apo‐form irrespective of the presence of cobalt in the medium. On the contrary, SCNase co‐expressed with P15K, encoded just downstream of SCNase genes, in cobalt‐enriched medium under the optimized condition (SCNase (+P15K) ) possessed 0.86 Co atom/αβγ trimer and exhibited 78% of the activity of native SCNase. SCNase (+P15K) showed a UV–Vis absorption peak characteristic of the SCNase cobalt center. About 70% of SCNase (+P15K) had the γCys131‐SO 2 H modification. These results indicate that SCNase (+P15K) is the active holo‐SCNase. P15K is likely to promote the functional expression of SCNase probably by assisting the incorporation of cobalt ion.