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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 glycosyltransferase enzyme glycogenin (GYG) (ED Figure 1). Deficiencies in glycogen formation cause neurodegenerative and metabolic disease (ED Figure 1b). 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 catalyzing its own stepwise auto glucosylation to form a covalently-bound gluco-oligosaccharide chain at initiation site Tyr195. To date, an inability to access homogeneous glycoforms of this protein, which unusually acts as both catalyst and substrate, has precluded precise mechanistic studies. 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 tri-phasic kinetics and substrate plasticity in GYG’s use of sugar substrates. This reveals a tolerant but ‘proof-read’ mechanism that underlies the precision of this vital metabolic process. This 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
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Raich, Lluis
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Gafitescu, Maria
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Machida, Takuya
42eeeac2-2937-4998-8228-a5803a50f198
Iglesias-Fernandez, Javier
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Lee, Seung Seo
ee34fa26-5fb6-48c8-80c2-1f13ec4ccceb
Spicer, Chris D.
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Rovira, Carme
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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
743305ec-326f-4049-900d-89bda7adedcf
Raich, Lluis
bb6dd93c-126c-48e3-9682-87de69762dcf
Gafitescu, Maria
0524cabc-16a5-4d85-8db8-8011d209baa8
Machida, Takuya
42eeeac2-2937-4998-8228-a5803a50f198
Iglesias-Fernandez, Javier
8eff1863-f9af-4600-82f5-6bcbc4584efe
Lee, Seung Seo
ee34fa26-5fb6-48c8-80c2-1f13ec4ccceb
Spicer, Chris 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, Raich, Lluis, Gafitescu, Maria, Machida, Takuya, Iglesias-Fernandez, Javier, Lee, Seung Seo, Spicer, Chris D., Rovira, Carme, Yue, Wyatt W. and Davis, Benjamin G. (2018) Palladium-mediated enzyme activation suggests multiphase initiation of glycogenesis. Nature, 563, 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 glycosyltransferase enzyme glycogenin (GYG) (ED Figure 1). Deficiencies in glycogen formation cause neurodegenerative and metabolic disease (ED Figure 1b). 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 catalyzing its own stepwise auto glucosylation to form a covalently-bound gluco-oligosaccharide chain at initiation site Tyr195. To date, an inability to access homogeneous glycoforms of this protein, which unusually acts as both catalyst and substrate, has precluded precise mechanistic studies. 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 tri-phasic kinetics and substrate plasticity in GYG’s use of sugar substrates. This reveals a tolerant but ‘proof-read’ mechanism that underlies the precision of this vital metabolic process. This 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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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: https://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: 24 Apr 2019 04:01

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