Two-substrate association with the 20S proteasome at single-molecule level
AUTOR(ES)
Hutschenreiter, Silke
FONTE
Nature Publishing Group
RESUMO
The bipartite structure of the proteasome raises the question of functional significance. A rational design for unraveling mechanistic details of the highly symmetrical degradation machinery from Thermoplasma acidophilum pursues orientated immobilization at metal-chelating interfaces via affinity tags fused either around the pore apertures or at the sides. End-on immobilization of the proteasome demonstrates that one pore is sufficient for substrate entry and product release. Remarkably, a ‘dead-end' proteasome can process only one substrate at a time. In contrast, the side-on immobilized and free proteasome can bind two substrates, presumably one in each antechamber, with positive cooperativity as analyzed by surface plasmon resonance and single-molecule cross-correlation spectroscopy. Thus, the two-stroke engine offers the advantage of speeding up degradation without enhancing complexity.
