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Affinity purification of the Legionella effector SidL in HEK293T cells

ABSTRACT: Legionella pneumophila is an intracellular bacterial pathogen that encodes almost 300 effector proteins, many of which target various aspects of its eukaryotic host. While the physiological roles of many effectors are not understood, the focused study of individual effectors provides insight into their often novel and unique biochemistries. Here, we characterize the function of one effector, SidL, who was reported to inhibit host translation. We find that SidL blocks the nutrient-responsive translation regulator mTORC1, suggesting that translation inhibition is a downstream consequence of SidL activity. We use computational analyses that classify SidL as a member of a phosphotransferase and oxidoreductase superfamily, including a recently described protein-modifying adenylyltransferase; however, SidL does not adenylate proteins. Instead we use in vitro biochemistry to discover that SidL adenylates the glycolysis intermediate 3-phosphoglycerate (3PG), producing the novel metabolite 2-AMP-3PG. In cells, SidL disrupts glycolysis with a notable loss of 3PG and downstream intermediates. We propose that SidL expression consumes 3PG, causing metabolic perturbations that inactivate mTORC1, consequentially inhibiting translation. To our knowledge, this is the first report of a bacterial effector uniquely modifying an intermediate in a conserved metabolic pathway and suggests a novel mechanism by which bacteria modulate host metabolism.

INSTRUMENT(S):

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Hek-293t Cell, Cell Culture

SUBMITTER: Joshua Black

LAB HEAD: Rachel Green

PROVIDER: PXD068090 | Pride | 2026-07-24

REPOSITORIES: Pride

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			Action	DRS
	Baits_noFLAGtag.fasta	Fasta
	JS-CS-E482_240610_GreenR_JB_S1_DIA_10pct.raw	Raw
	JS-CS-E482_240610_GreenR_JB_S2_DIA_10pct.raw	Raw
	JS-CS-E482_240610_GreenR_JB_S3_DIA_10pct.raw	Raw
	JS-CS-E482_240610_GreenR_JB_S4_DIA_10pct.raw	Raw

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Affinity purification of the Legionella effector SidL in HEK293T cells

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