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Palladium-mediated enzyme activation suggests multiphase initiation of glycogenesis

Palladium-mediated enzyme activation suggests multiphase initiation of glycogenesis
Palladium-mediated enzyme activation suggests multiphase initiation of glycogenesis
Biosynthesis of glycogen, the essential glucose (and hence energy) storage molecule in humans, animals and fungi, is initiated by the glycosyltransferase enzyme, glycogenin (GYG). Deficiencies in glycogen formation cause neurodegenerative and metabolic disease, and mouse knockout and inherited human mutations of GYG impair glycogen synthesis. GYG acts as a ‘seed core’ for the formation of the glycogen particle by catalysing its own stepwise autoglucosylation to form a covalently bound gluco-oligosaccharide chain at initiation site Tyr 195. Precise mechanistic studies have so far been prevented by an inability to access homogeneous glycoforms of this protein, which unusually acts as both catalyst and substrate.
Here we show that unprecedented direct access to different, homogeneously glucosylated states of GYG can be accomplished through a palladium-mediated enzyme activation ‘shunt’ process using on-protein C–C bond formation. Careful mimicry of GYG intermediates recapitulates catalytic activity at distinct stages, which in turn allows discovery of triphasic kinetics and substrate plasticity in GYG’s use of sugar substrates. This reveals a tolerant but ‘proofread’ mechanism that underlies the precision of this metabolic process. The present demonstration of direct, chemically controlled access to intermediate states of active enzymes suggests that such ligation-dependent activation could be a powerful tool in the study of mechanism.
0028-0836
235–240
Bilyard, Matthew K.
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Bailey, Henry J.
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Raich, Lluís
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Gafitescu, Maria A.
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Machida, Takuya
42eeeac2-2937-4998-8228-a5803a50f198
Iglésias-Fernández, Javier
8eff1863-f9af-4600-82f5-6bcbc4584efe
Lee, Seung Seo
ee34fa26-5fb6-48c8-80c2-1f13ec4ccceb
Spicer, Christopher D.
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Rovira, Carme
0cfae9b2-fbf5-4127-83c9-995ccd46f25d
Yue, Wyatt W.
5151445e-a83d-4835-9535-4d5d91e8a085
Davis, Benjamin G.
64b1001d-1a32-404c-b947-6c4bcd35df9f
Bilyard, Matthew K.
513071c3-1ac5-421b-8156-ab1946e620d5
Bailey, Henry J.
743305ec-326f-4049-900d-89bda7adedcf
Raich, Lluís
bb6dd93c-126c-48e3-9682-87de69762dcf
Gafitescu, Maria A.
0524cabc-16a5-4d85-8db8-8011d209baa8
Machida, Takuya
42eeeac2-2937-4998-8228-a5803a50f198
Iglésias-Fernández, Javier
8eff1863-f9af-4600-82f5-6bcbc4584efe
Lee, Seung Seo
ee34fa26-5fb6-48c8-80c2-1f13ec4ccceb
Spicer, Christopher D.
69a3a83a-337e-489a-a6ee-d0352bce8601
Rovira, Carme
0cfae9b2-fbf5-4127-83c9-995ccd46f25d
Yue, Wyatt W.
5151445e-a83d-4835-9535-4d5d91e8a085
Davis, Benjamin G.
64b1001d-1a32-404c-b947-6c4bcd35df9f

Bilyard, Matthew K., Bailey, Henry J., Raich, Lluís, Gafitescu, Maria A., Machida, Takuya, Iglésias-Fernández, Javier, Lee, Seung Seo, Spicer, Christopher D., Rovira, Carme, Yue, Wyatt W. and Davis, Benjamin G. (2018) Palladium-mediated enzyme activation suggests multiphase initiation of glycogenesis. Nature, 563 (7730), 235–240. (doi:10.1038/s41586-018-0644-7).

Record type: Article

Abstract

Biosynthesis of glycogen, the essential glucose (and hence energy) storage molecule in humans, animals and fungi, is initiated by the glycosyltransferase enzyme, glycogenin (GYG). Deficiencies in glycogen formation cause neurodegenerative and metabolic disease, and mouse knockout and inherited human mutations of GYG impair glycogen synthesis. GYG acts as a ‘seed core’ for the formation of the glycogen particle by catalysing its own stepwise autoglucosylation to form a covalently bound gluco-oligosaccharide chain at initiation site Tyr 195. Precise mechanistic studies have so far been prevented by an inability to access homogeneous glycoforms of this protein, which unusually acts as both catalyst and substrate.
Here we show that unprecedented direct access to different, homogeneously glucosylated states of GYG can be accomplished through a palladium-mediated enzyme activation ‘shunt’ process using on-protein C–C bond formation. Careful mimicry of GYG intermediates recapitulates catalytic activity at distinct stages, which in turn allows discovery of triphasic kinetics and substrate plasticity in GYG’s use of sugar substrates. This reveals a tolerant but ‘proofread’ mechanism that underlies the precision of this metabolic process. The present demonstration of direct, chemically controlled access to intermediate states of active enzymes suggests that such ligation-dependent activation could be a powerful tool in the study of mechanism.

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316059 2 merged 1535644194 - Accepted Manuscript
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Accepted/In Press date: 31 August 2018
e-pub ahead of print date: 24 October 2018
Published date: 8 November 2018
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Identifiers

Local EPrints ID: 425590
URI: http://eprints.soton.ac.uk/id/eprint/425590
ISSN: 0028-0836
PURE UUID: 3e694440-1800-4a99-a32d-c76cff0aaaed
ORCID for Seung Seo Lee: ORCID iD orcid.org/0000-0002-8598-3303

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Date deposited: 25 Oct 2018 16:30
Last modified: 11 Aug 2020 04:01

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