Cells constantly use and recycle their proteins in a process called protein quality control (PQC). Imbalance of PQC leads to accumulation of dysfunctional proteins, thus leading to a variety of diseases. Hsp70 is a central mediator of PQC by collaborating with J-domain proteins (JDPs) to regulate client protein folding and degradation. Despite the importance of this system, the molecular mechanisms governing these interactions remain unclear. This study employs computationally designed proteins to create synthetic Hsp70-binding proteins that can either enhance or inhibit its activity. One of these designs mimic native JDPs in promoting refolding of denatured proteins and can modulate intracellular condensates, revealing insights into the roles of these condensates. This work provides insight in three aspects: (1) computational and experimental methods to design synthetic binding proteins, (2) advances our understanding of Hsp70 interactions that regulate PQC, (3) introduces modular tools to manipulate Hsp70 activity and condensates. We anticipate that these results and tools will accelerate our understanding of PQC and manipulation of PQC in diseases.
