ArticleBio-protocol2024
Real-Time Monitoring of ATG8 Lipidation in vitro Using Fluorescence Spectroscopy.
Article in Bio-protocol, 2024. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 1 paper.
What it found
Each row is one number read from the abstract, on the scale the paper reported it, with its interval. Left of the dashed line favours the treatment, right favours the comparator. Under each row is the sentence it came from. New to these charts? A ten-minute tutorial.
The abstract states no effect estimate the extractor could read, or names no intervention and outcome on the map, so this paper lights no cell and moves no belief. It is still indexed, cited and linked below.
The trial behind it
Trials whose registry record cites this paper, or whose number appears in the abstract. A trial that started after this paper was published is citing it as background, not reporting it.
Neither the registry nor the abstract names a trial number. If this is a trial report, that itself is worth knowing.
Who cites it
1 citing paper in PubMed, 2 citations in OpenAlex.
- Review
Corrections and comments
PubMed lists nothing against this paper. Absence here is not a guarantee, only a check that was made.
Authors and funding
3 authors at 2 institutions in 2 countries.
Funding
Abstract
Autophagy is an essential catabolic pathway used to sequester and engulf cytosolic substrates via a unique double-membrane structure, called an autophagosome. The ubiquitin-like ATG8 proteins play an important role in mediating autophagosome membrane expansion. They are covalently conjugated to phosphatidylethanolamine (PE) on the autophagosomes via a ubiquitin-like conjugation system called ATG8 lipidation. In vitro reconstitution of ATG8 lipidation with synthetic liposomes has been previously established and used widely to characterise the function of the E1 ATG7, the E2 ATG3, and the E3 complex ATG12-ATG5-ATG16L1. However, there is still a lack of a tool to provide kinetic measurements of this enzymatic reaction. In this protocol, we describe a real-time lipidation assay using NBD-labelled ATG8. This real-time assay can distinguish the formation of ATG8 intermediates (ATG7~ATG8 and/or ATG3~ATG8) and, finally, ATG8-PE conjugation. It allows kinetic characterisation of the activity of ATG7, ATG3, and the E3 complex during ATG8 lipidation. Furthermore, this protocol can be adapted to characterise the upstream regulators that may affect protein activity in ATG8 lipidation reaction with a kinetic readout. Key features • Preparation of ATG7 E1 from insect cells (Sf9 cells). • Preparation of ATG3 E2 from bacteria (
Indexed as
Identifiers
What OpenQuestion holds
Registered trials
Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.