# Engineering Artificial Mitochondria with Self‐Amplifying Proton Generation for Autonomous Energy Supply and Metabolic Coupling in Artificial Cells

## TL;DR

Researchers created artificial mitochondria that generate energy autonomously in artificial cells, mimicking key features of real mitochondria.

## Contribution

A self-amplifying proton gradient system using co-compartmentalized enzymes in nanocapsules for autonomous ATP production in artificial cells.

## Key findings

- The nanocapsules enable a self-reinforcing enzymatic cascade that amplifies proton production.
- Artificial mitochondria integrated into GUVs support glucose-powered ATP generation and NADH biosynthesis.
- The system improves enzyme stability and eliminates H2O2 toxicity through co-compartmentalization.

## Abstract

A continuous and autonomous energy supply is essential for sustaining life‐like biochemical processes in artificial cells. Although considerable efforts have been devoted to engineering artificial organelles that emulate mitochondrial energy conversion, the generation of a robust transmembrane proton gradient—essential for driving efficient ATP production—remains a major challenge. Here, we present a mitochondria‐mimicking ATP nano‐generator constructed through quantitative co‐compartmentalization of glucose oxidase and catalase within silica nanocapsules. Enzymes are encapsulated in situ during the formation of core‐shell nanocapsules, enabling precise loading, effective protection, and creation of a confined nanoscale reaction chamber that fosters catalytic synergy. Within this microenvironment, catalase rapidly decomposes H2O2 to generate O2, which is in turn utilized by glucose oxidase—thus establishing a self‐reinforcing enzymatic cascade that amplifies proton production. After coating the enzyme‐loaded nanocapsules with an ATPase‐integrated liposome bilayer to construct the artificial mitochondrion, the resulting proton gradient across the membrane efficiently drives ATP synthase rotation, enabling high‐yield ATP production. When integrated into giant unilamellar vesicles (GUVs) as synthetic cell models, this system supports autonomous nicotinamide adenine dinucleotide (NADH) biosynthesis and glucose‐powered oxidative phosphorylation, mimicking key metabolic features of living mitochondria. This work establishes an effective and versatile platform for engineering energy‐autonomous artificial living systems, advancing the state of the art of bottom‐up synthetic biology.

A mitochondria‐mimicking ATP generator improves the enzyme instability of enzyme‐based artificial mitochondria and eliminates the toxicity of H2O2 via quantitative enzyme co‐compartmentalization. The resulting artificial mitochondria can establish a sustained proton gradient for ATP synthesis. The integration of artificial mitochondria into artificial cells can enable glucose‐fueled ATP generation, powering an autonomous NADH biosynthesis.