Biology subjects

Pardo-Hernandez, C.

Publications and source records attributed to Pardo-Hernandez, C..

2 recordsLinked to original sources

bioRxiv2021

A dual allosteric pathway drives human mitochondrial Lon

The hexameric, barrel-forming, AAA+ protease Lon is critical for maintaining mitochondrial matrix protein homeostasis. Efficient substrate processing by Lon requires the coordinated action of six protomers. Despite Lons importance for human health, the molecular bases for Lons substrate recognition and processing remain unclear. Here, we use a combination of biochemistry and electron cryomicroscopy (cryo-EM) to unveil the structural and functional basis for full-length human mitochondrial Lons degradation of mitochondrial transcription factor A (TFAM). We show how opposing protomers in the Lon hexamer barrel interact through their N-terminal domains to give what resembles three feet above the barrel and help to form a triangular pore located just above the entry pore to the barrel. The interactions between opposing protomers constitute a primary allosteric regulation of Lon activity. A secondary allosteric regulation consists of an inter-subunit signaling element in the ATPase domains. By considering the ATP or ADP load in each protomer, we show how this dual allosteric mechanism in Lon achieves coordinated ATP hydrolysis and substrate processing. This mechanism enforces sequential anti-clockwise ATP hydrolysis resulting in a coordinated hand-over-hand translocation of the substrate towards the protease active sites.

biochemistry↗

bioRxiv2020

In vivo and in vitro mechanistic characterization of a clinically relevant PolγA mutation

Mutations in POLG, encoding POL{gamma}A, the catalytic subunit of the mitochondrial DNA polymerase, cause a spectrum of disorders characterized by mtDNA instability. However, the molecular pathogenesis of POLG-related diseases is poorly understood and efficient treatments are missing. Here, we generated a POLGA449T/A449T mouse model, which reproduces the most common human recessive mutation of POLG, encoding the A467T change, and dissected the mechanisms underlying pathogenicity. We show that the A449T mutation impairs DNA binding and mtDNA synthesis activities of POL{gamma} in vivo and in vitro. Interestingly, the A467T mutation also strongly impairs interactions with POL{gamma}B, the homodimeric accessory subunit of holo-POL{gamma}. This allows the free POL{gamma}A to become a substrate for LONP1 protease degradation, leading to dramatically reduced levels of POL{gamma}A, which in turn exacerbates the molecular phenotypes of PolgA449T/A449T mice. Importantly, we validated this mechanism for other mutations affecting the interaction between the two POL{gamma} subunits. We suggest that LONP1 dependent degradation of POL{gamma}A can be exploited as a target for the development of future therapies.

genetics↗