Crystal structure of a low molecular weight activator Blm-pep with yeast 20S proteasome – insights into the enzyme activation mechanism

Abstract

Proteasomes are responsible for protein turnover in eukaryotic cells, degrading short-lived species but also removing improperly folded or oxidatively damaged ones. Dysfunction of a proteasome results in gradual accumulation of misfolded/damaged proteins, leading to their aggregation. It has been postulated that proteasome activators may facilitate removal of such aggregation-prone proteins and thus prevent development of neurodegenerative disorders. However, the discovery of pharmacologically relevant compounds is hindered by insufficient structural understanding of the activation process. In this study we provide a model peptidic activator of human proteasome and analyze the structure-activity relationship within this novel scaffold. The binding mode of the activator at the relevant pocket within the proteasome has been determined by X-ray crystallography. This crystal structure provides an important basis for rational design of pharmacological compounds. Moreover, by providing a novel insight into the proteasome gating mechanism, our results allow the commonly accepted model of proteasome regulation to be revisited.

Pyrazolones Activate the Proteasome by Gating Mechanisms and Protect Neuronal Cells from β‐Amyloid Toxicity

“…Moreover, the 19S particle, as well as other 20S activators, induces complex conformational changes that allosterically facilitates substrate translocation and hydrolysis in the catalytic β‐subunits . All known types of RPs, including 19S, PA200 and PA28 include an HbYX (hydrophobic, Hb; Tyrosine, Y; any amino acid, X) motif which plays a key role in steering RPs/20S assembly and in promoting gate opening . For this reason, many peptides harnessing the HbYX motif including i) the activating loop and C‐terminus of PA28, the cathelicidin antibacterial peptide PR39, the octa peptide Arg(8) and the HIV‐1 Tat protein, were shown to be allosteric 20S activators.…”

New Peptide-Based Pharmacophore Activates 20S Proteasome

“…The inter-subunit pockets are used to attach regulatory proteins: PA28/REG (all pockets), PA200 (proteasome activator of 200 kDa; α5/α6 pocket), as well as the Rpt subunits of the 19S particle (all pockets except α6/α7 and α7/α1) [26,36,40]. Peptide-activators of the core that utilize the structure of docking fragments of these natural activators were found to bind into the α5/α6 pocket, while a small molecule activator TCH-165 reportedly preferred the α1/α2 site [19,20,41]. Binding of all these ligands resulted in opening or at least destabilizing the gate in the center of the α face, as revealed by crystal structures, cryoEM (cryo-electron microscopy) and atomic force microscopy (AFM) imaging [19,20,25,[40][41][42][43].…”

“…Peptide-activators of the core that utilize the structure of docking fragments of these natural activators were found to bind into the α5/α6 pocket, while a small molecule activator TCH-165 reportedly preferred the α1/α2 site [19,20,41]. Binding of all these ligands resulted in opening or at least destabilizing the gate in the center of the α face, as revealed by crystal structures, cryoEM (cryo-electron microscopy) and atomic force microscopy (AFM) imaging [19,20,25,[40][41][42][43]. Gate opening is prerequisite for the uptake of substrates and release of products from the concealed catalytic chamber of the core proteasome ( Figure 1A).…”

Small‐Molecule Inhibitors of the Proteasome's Regulatory Particle

“…It has long been hypothesized, for example, that increasing proteasome activity could combat premature aging or neurological diseases. There are several examples of molecules that can stimulate the activity of the proteasome . More recently, it has been proposed that proteasome inhibitors might aid in treating autoimmune diseases .…”