Dataset Information

Cullin-independent recognition of HHARI substrates by a dynamic RBR catalytic domain

ABSTRACT: RING-Between-RING (RBR) E3 ligases mediate ubiquitin transfer through an obligate E3- ubiquitin thioester intermediately prior to substrate ubiquitination. While RBRs share a conserved catalytic module, substrate recruitment mechanisms remain enigmatic and the relevant domains have yet to be identified for any member of the class. Here we characterize the interaction between the auto-inhibited RBR, HHARI (AriH1), and its target protein, 4EHP, using a combination of XL-MS, HDX-MS, NMR, and biochemical studies. The results show that 1) a di-aromatic surface on the catalytic HHARI Rcat domain forms a binding platform for substrates and 2) a phospho-mimetic mutation on the auto-inhibitory Ariadne domain of HHARI promotes release and reorientation of Rcat for transthiolation and substrate modification. The findings represent the first identification of a direct binding interaction between a RING-Between-RING ligase and its substrate.

INSTRUMENT(S): Q Exactive HF

ORGANISM(S): Homo Sapiens (human)

SUBMITTER: Alex Zelter

LAB HEAD: Rachel E. Klevit

PROVIDER: PXD030849 | Pride | 2023-05-24

REPOSITORIES: Pride

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Cullin-independent recognition of HHARI substrates by a dynamic RBR catalytic domain.

Reiter Katherine H KH Zelter Alex A Janowska Maria K MK Riffle Michael M Shulman Nicholas N MacLean Brendan X BX Tamura Kaipo K Chambers Matthew C MC MacCoss Michael J MJ Davis Trisha N TN Guttman Miklos M Brzovic Peter S PS Klevit Rachel E RE

Structure (London, England : 1993) 20220617 9

RING-between-RING (RBR) E3 ligases mediate ubiquitin transfer through an obligate E3-ubiquitin thioester intermediate prior to substrate ubiquitination. Although RBRs share a conserved catalytic module, substrate recruitment mechanisms remain enigmatic, and the relevant domains have yet to be identified for any member of the class. Here we characterize the interaction between the auto-inhibited RBR, HHARI (AriH1), and its target protein, 4EHP, using a combination of XL-MS, HDX-MS, NMR, and bioch ...[more]

PMID: 35716664

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			Action	DRS
	EH208a_raw.zip	Other
	EH208a_results.zip	Other
	EH208b_raw.zip	Other
	EH208b_results.zip	Other
	EH208c_raw.zip	Other