Loop-mediated regulation and base flipping drive RNA cleavage by human mitochondrial PNPase

Area of Science:

• Biochemistry
• Molecular Biology
• Structural Biology

Background:

• Human polynucleotide phosphorylase (hPNPase) is a trimeric exoribonuclease essential for mitochondrial RNA metabolism.
• Mutations in hPNPase are associated with mitochondrial diseases, highlighting its role in RNA homeostasis.

Purpose of the Study:

• To elucidate the molecular basis of hPNPase's catalytic mechanism.
• To understand the structural consequences of active-site mutations in hPNPase.

Main Methods:

• High-resolution electron cryo-microscopy (cryo-EM) was used to visualize hPNPase.
• Three distinct functional states during RNA degradation were captured: loading, pre-catalytic, and catalytic.

Main Results:

• Flexible loops in the loading state facilitate substrate RNA recruitment.
• In the pre-catalytic state, Mg2+ stabilizes nucleotide reorientation for cleavage.
• The catalytic state shows a nucleophilic attack on the RNA backbone mediated by active-site residues.

Conclusions:

• The study provides a biochemical framework for hPNPase-mediated RNA turnover.
• The findings clarify the catalytic mechanism of hPNPase.
• Active-site integrity is demonstrated as crucial for efficient RNA degradation by hPNPase.