title: Protein unfolding by a AAA+ protease is dependent on ATP-hydrolysis rates and substrate energy landscapes.
abstract: In the AAA+ ClpXP protease, repetitive cycles of ATP hydrolysis by ClpX unfold ssrA-tagged substrates, which are unraveled vectorially starting at the C-terminal tag and translocated into ClpP for degradation. When the ATP-hydrolysis rate drops below a critical threshold, ClpXP fails to degrade ssrA-tagged green fluorescent protein (GFP) but degrades other tagged proteins, including some that are more stable. Our results support a model in which ClpX unfolding of GFP progresses via a metastable short-lived intermediate, which must be captured by several fast ATP-dependent translocation steps to prevent the protein from refolding and therefore escaping degradation. Thus, AAA+ proteases may be more or less suited to degradation of specific substrates depending on their ability to disrupt native structure and to trap partially unfolded intermediates successfully. We propose that cellular conditions or adaptors that alter ATP-hydrolysis rates could control this trapping activity of AAA+ enzymes.
authors.affiliation: Department of Biology, 77 Massachusetts Avenue, Cambridge, Massachusetts 02139, USA.
authors.forename: Andreas
authors.initials: A
authors.lastname: Martin
authors.forename: Tania A
authors.initials: TA
authors.lastname: Baker
authors.forename: Robert T
authors.initials: RT
authors.lastname: Sauer
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism*; D049071:Endopeptidase Clp / Q000737:chemistry* / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000378:metabolism*; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006868:Hydrolysis; D007700:Kinetics; D008954:Models, Biological; D008958:Models, Molecular; D017510:Protein Folding; D013997:Time Factors
chemical_list: D029968:Escherichia coli Proteins; D049452:Green Fluorescent Proteins; D000255:Adenosine Triphosphate; C483573:ClpXP protease, E coli; D049071:Endopeptidase Clp