Pyrococcus horikoshii TET2 Peptidase Assembling Process and Associated Functional Regulation

Type Article
Date 2013-08
Language English
Author(s) Appolaire Alexandre1, 2, 3, Rosenbaum Eva1, 2, 3, Dura M. Asuncion1, 2, 3, Colombo Matteo1, 2, 3, Marty Vincent1, 2, 3, Savoye Marjolaine Noirclerc1, 2, 3, Godfroy AnneORCID4, Schoehn Guy1, 2, 3, Girard Eric1, 2, 3, Gabel Frank1, 2, 3, Franzetti Bruno1, 2, 3
Affiliation(s) 1 : CNRS, Inst Biol Struct, UMR5075, F-38027 Grenoble, France.
2 : CEA Grenoble, F-38054 Grenoble, France.
3 : Univ Grenoble 1, F-38027 Grenoble, France.
4 : IFREMER, UMR6197, Lab Microbiol Environm Extremes, F-29280 Plouzane, France.
Source Journal Of Biological Chemistry (0021-9258) (Amer Soc Biochemistry Molecular Biology Inc), 2013-08 , Vol. 288 , N. 31 , P. 22542-22554
DOI 10.1074/jbc.M113.450189
WOS© Times Cited 11
Abstract Tetrahedral (TET) aminopeptidases are large polypeptide destruction machines present in prokaryotes and eukaryotes. Here, the rules governing their assembly into hollow 12-subunit tetrahedrons are addressed by using TET2 from Pyrococcus horikoshii (PhTET2) as a model. Point mutations allowed the capture of a stable, catalytically active precursor. Small angle x- ray scattering revealed that it is a dimer whose architecture in solution is identical to that determined by x- ray crystallography within the fully assembled TET particle. Small angle x- ray scattering also showed that the reconstituted PhTET2 dodecameric particle displayed the same quaternary structure and thermal stability as the wild-type complex. The PhTET2 assembly intermediates were characterized by analytical ultracentrifugation, native gel electrophoresis, and electron microscopy. They revealed that PhTET2 assembling is a highly ordered process in which hexamers represent the main intermediate. Peptide degradation assays demonstrated that oligomerization triggers the activity of the TET enzyme toward large polypeptidic substrates. Fractionation experiments in Pyrococcus and Halobacterium cells revealed that, in vivo, the dimeric precursor co-exists together with assembled TET complexes. Taken together, our observations explain the biological significance of TET oligomerization and suggest the existence of a functional regulation of the dimer-dodecamer equilibrium in vivo.
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Appolaire Alexandre, Rosenbaum Eva, Dura M. Asuncion, Colombo Matteo, Marty Vincent, Savoye Marjolaine Noirclerc, Godfroy Anne, Schoehn Guy, Girard Eric, Gabel Frank, Franzetti Bruno (2013). Pyrococcus horikoshii TET2 Peptidase Assembling Process and Associated Functional Regulation. Journal Of Biological Chemistry, 288(31), 22542-22554. Publisher's official version : , Open Access version :