Trapping conformational states along ligand-binding dynamics of peptide deformylase: the impact of induced fit on enzyme catalysis.
Bottom Line: Ligand-induced reshaping of a hydrophobic pocket drives closure of the active site, which is finally "zipped up" by additional binding interactions.Together with biochemical analyses, these data allow a coherent reconstruction of the sequence of events leading from the encounter complex to the key-lock binding state of the enzyme.A "movie" that reconstructs this entire process can be further extrapolated to catalysis.
Affiliation: CNRS, ISV, UPR2355, Gif-sur-Yvette, France.
For several decades, molecular recognition has been considered one of the most fundamental processes in biochemistry. For enzymes, substrate binding is often coupled to conformational changes that alter the local environment of the active site to align the reactive groups for efficient catalysis and to reach the transition state. Adaptive substrate recognition is a well-known concept; however, it has been poorly characterized at a structural level because of its dynamic nature. Here, we provide a detailed mechanism for an induced-fit process at atomic resolution. We take advantage of a slow, tight binding inhibitor-enzyme system, actinonin-peptide deformylase. Crystal structures of the initial open state and final closed state were solved, as well as those of several intermediate mimics captured during the process. Ligand-induced reshaping of a hydrophobic pocket drives closure of the active site, which is finally "zipped up" by additional binding interactions. Together with biochemical analyses, these data allow a coherent reconstruction of the sequence of events leading from the encounter complex to the key-lock binding state of the enzyme. A "movie" that reconstructs this entire process can be further extrapolated to catalysis.
Related in: MedlinePlus
Mentions: We investigated the existence of lowly populated, alternative conformations of apoPDF. To probe the occurrence of alternate conformers in the crystalline state of PDF, the new Ringer program is the most suitable investigation tool ,. Ringer searches for evidence of alternate rotamers by systematically sampling electron density maps—free of model bias—around the dihedral angles of protein side chains. Two independent WT open datasets of the apoenzyme, including a high-resolution set (1.3 Å), were used in the analysis. Ringer analysis revealed the existence of only one rotamer of most side chains of either molecule in the asymmetric unit, including the three main residues primarily involved in conformation change—that is, Ile 42, Phe58, and Ile130 (Figure 4A). Ringer analysis showed evidence for unmodeled alternate conformers for very few residues, including Ile121 and Phe87, or Phe119 to a much lesser extent (Figure S7). There is therefore no evidence for the occurrence of a closed conformation in the apostructure of AtPDF, supporting the hypothesis that the conformational change was essentially induced by the binding of actinonin rather than from conformational selection among multiple states occurring in the crystalline state.