Discovery of posttranslational maturation by self-subunit swapping.
Several general mechanisms of metallocenter biosynthesis have been reported and reviewed, and in all cases, the components or subunits of an apoprotein remain in the final holoprotein. Here, we first discovered that one subunit of an apoenzyme did not remain in the functional holoenzyme. The cobalt-containing low-molecular-mass nitrile hydratase (L-NHase) of Rhodococcus rhodochrous J1 consists of beta- and alpha-subunits encoded by the nhlBA genes, respectively. An ORF, nhlE, just downstream of nhlBA, was found to be necessary for L-NHase activation. In contrast to the cobalt-containing L-NHase (holo-L-NHase containing Cys-SO(2)(-) and Cys-SO(-) metal ligands) derived from nhlBAE, the gene products derived from nhlBA were cobalt-free L-NHase (apo-L-NHase lacking oxidized cysteine residues). We discovered an L-NHase maturation mediator, NhlAE, consisting of NhlE and the cobalt- and oxidized cysteine-containing alpha-subunit of L-NHase. The incorporation of cobalt into L-NHase was shown to depend on the exchange of the nonmodified cobalt-free alpha-subunit of apo-L-NHase with the cobalt-containing cysteine-modified alpha-subunit of NhlAE. This is a posttranslational maturation process different from general mechanisms of metallocenter biosynthesis known so far: the unexpected behavior of a protein in a protein complex, which we named "self-subunit swapping."
Zhou
Zhemin
Z
Institute of Applied Biochemistry and Graduate School of Life and Environmental Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki 305-8572, Japan.
Hashimoto
Yoshiteru
Y
Shiraki
Kentaro
K
Kobayashi
Michihiko
M
D001051:Apoenzymes / Q000235:genetics / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D003035:Cobalt / Q000378:metabolism*; D003545:Cysteine / Q000378:metabolism; D004789:Enzyme Activation; D020035:Holoenzymes / Q000235:genetics / Q000378:metabolism; D006836:Hydro-Lyases / Q000235:genetics / Q000378:metabolism*; D008667:Metalloproteins / Q000235:genetics / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D021122:Protein Subunits / Q000235:genetics / Q000378:metabolism; D012240:Rhodococcus / Q000201:enzymology* / Q000235:genetics
D001051:Apoenzymes; D001426:Bacterial Proteins; D020035:Holoenzymes; D008667:Metalloproteins; D021122:Protein Subunits; D003035:Cobalt; D006836:Hydro-Lyases; C048831:nitrile hydratase; D003545:Cysteine