Structural analysis reveals a pyruvate-binding activator site in the Agrobacterium tumefaciens ADP–glucose pyrophosphorylase
Abstract
The pathways for biosynthesis of glycogen in bacteria and starch in plants are evolutionarily and biochemically related. They are regulated primarily by ADP–glucose pyrophosphorylase, which evolved to satisfy metabolic requirements of a particular organism. Despite the importance of these two pathways, little is known about the mechanism that controls pyrophosphorylase activity or the location of its allosteric sites. Here, in this work, we report pyruvate-bound crystal structures of ADP–glucose pyrophosphorylase from the bacterium Agrobacterium tumefaciens, identifying a previously elusive activator site for the enzyme. We found that the tetrameric enzyme binds two molecules of pyruvate in a planar conformation. Each binding site is located in a crevice between the C-terminal domains of two subunits where they stack via a distinct β-helix region. Pyruvate interacts with the side chain of Lys-43 and with the peptide backbone of Ser-328 and Gly-329 from both subunits. These structural insights led to the design of two variants with altered regulatory properties. In one variant (K43A), the allosteric effect was absent, whereas in the other (G329D), the introduced Asp mimicked the presence of pyruvate. The latter generated an enzyme that was preactivated and insensitive to further activation by pyruvate. Our study furnishes a deeper understanding ofmore »
- Authors:
-
[1];
- Loyola Univ., Chicago, IL (United States)
- Loyola Univ., Chicago, IL (United States); Univ. Nacional del Litoral-Consejo Nacional de Investigaciones Científicas y Técnicas (UNL-CONICET), Santa Fe (Argentina)
- Univ. Nacional del Litoral-Consejo Nacional de Investigaciones Científicas y Técnicas (UNL-CONICET), Santa Fe (Argentina)
- Publication Date:
- Research Org.:
- Argonne National Laboratory (ANL), Argonne, IL (United States). Advanced Photon Source (APS)
- Sponsoring Org.:
- USDOE Office of Science (SC), Biological and Environmental Research (BER); National Science Foundation (NSF); Agencia Nacional de Promoción Científica y Tecnológica (ANPCyT)
- OSTI Identifier:
- 1511729
- Grant/Contract Number:
- AC02-06CH11357; MCB 1616851; PICTs-2014-3362; PICTs-2015-0634; PICTs-2015-1767 PICTs-2014-3256
- Resource Type:
- Accepted Manuscript
- Journal Name:
- Journal of Biological Chemistry
- Additional Journal Information:
- Journal Volume: 294; Journal Issue: 4; Journal ID: ISSN 0021-9258
- Publisher:
- American Society for Biochemistry and Molecular Biology
- Country of Publication:
- United States
- Language:
- ENGLISH
- Subject:
- 59 BASIC BIOLOGICAL SCIENCES; glycogen; allosteric regulation; glucose; enzyme structure; pyruvate; allosterism; glucose-1-phosphate adenylyltransferase; glycogen biosynthesis; starch biosynthesis; enzyme evolution; pyrophosphorylase; polyglucan synthesis
Citation Formats
Hill, Benjamin L., Mascarenhas, Romila, Patel, Hiral P., Asencion Diez, Matías D., Wu, Rui, Iglesias, Alberto A., Liu, Dali, and Ballicora, Miguel A. Structural analysis reveals a pyruvate-binding activator site in the Agrobacterium tumefaciens ADP–glucose pyrophosphorylase. United States: N. p., 2018.
Web. doi:10.1074/jbc.RA118.004246.
Hill, Benjamin L., Mascarenhas, Romila, Patel, Hiral P., Asencion Diez, Matías D., Wu, Rui, Iglesias, Alberto A., Liu, Dali, & Ballicora, Miguel A. Structural analysis reveals a pyruvate-binding activator site in the Agrobacterium tumefaciens ADP–glucose pyrophosphorylase. United States. https://doi.org/10.1074/jbc.RA118.004246
Hill, Benjamin L., Mascarenhas, Romila, Patel, Hiral P., Asencion Diez, Matías D., Wu, Rui, Iglesias, Alberto A., Liu, Dali, and Ballicora, Miguel A. Tue .
"Structural analysis reveals a pyruvate-binding activator site in the Agrobacterium tumefaciens ADP–glucose pyrophosphorylase". United States. https://doi.org/10.1074/jbc.RA118.004246. https://www.osti.gov/servlets/purl/1511729.
@article{osti_1511729,
title = {Structural analysis reveals a pyruvate-binding activator site in the Agrobacterium tumefaciens ADP–glucose pyrophosphorylase},
author = {Hill, Benjamin L. and Mascarenhas, Romila and Patel, Hiral P. and Asencion Diez, Matías D. and Wu, Rui and Iglesias, Alberto A. and Liu, Dali and Ballicora, Miguel A.},
abstractNote = {The pathways for biosynthesis of glycogen in bacteria and starch in plants are evolutionarily and biochemically related. They are regulated primarily by ADP–glucose pyrophosphorylase, which evolved to satisfy metabolic requirements of a particular organism. Despite the importance of these two pathways, little is known about the mechanism that controls pyrophosphorylase activity or the location of its allosteric sites. Here, in this work, we report pyruvate-bound crystal structures of ADP–glucose pyrophosphorylase from the bacterium Agrobacterium tumefaciens, identifying a previously elusive activator site for the enzyme. We found that the tetrameric enzyme binds two molecules of pyruvate in a planar conformation. Each binding site is located in a crevice between the C-terminal domains of two subunits where they stack via a distinct β-helix region. Pyruvate interacts with the side chain of Lys-43 and with the peptide backbone of Ser-328 and Gly-329 from both subunits. These structural insights led to the design of two variants with altered regulatory properties. In one variant (K43A), the allosteric effect was absent, whereas in the other (G329D), the introduced Asp mimicked the presence of pyruvate. The latter generated an enzyme that was preactivated and insensitive to further activation by pyruvate. Our study furnishes a deeper understanding of how glycogen biosynthesis is regulated in bacteria and the mechanism by which transgenic plants increased their starch production. These insights will facilitate rational approaches to enzyme engineering for starch production in crops of agricultural interest and will promote further study of allosteric signal transmission and molecular evolution in this important enzyme family.},
doi = {10.1074/jbc.RA118.004246},
journal = {Journal of Biological Chemistry},
number = 4,
volume = 294,
place = {United States},
year = {Tue Nov 06 00:00:00 EST 2018},
month = {Tue Nov 06 00:00:00 EST 2018}
}
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Works referencing / citing this record:
Structural basis of glycogen metabolism in bacteria
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