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Title: Intrinsic disorder in the regulatory N-terminal domain of diacylglycerol acyltransferase 1 from Brassica napus

Abstract

Proteins with multifunctional regulatory domains often demonstrate structural plasticity or protein disorder, allowing the binding of multiple regulatory factors and post-translational modifications. While the importance of protein disorder is clear, it also poses a challenge for in vitro characterization. Here, we report protein intrinsic disorder in a plant molecular system, which despite its prevalence is less studied. We present a detailed biophysical characterization of the entire cytoplasmic N-terminal domain of Brassica napus diacylglycerol acyltransferase, (DGAT1), which includes an inhibitory module and allosteric binding sites. Our results demonstrate that the monomeric N-terminal domain can be stabilized for biophysical characterization and is largely intrinsically disordered in solution. This domain interacts with allosteric modulators of DGAT1, CoA and oleoyl-CoA, at micromolar concentrations. Furthermore, while solution scattering studies indicate conformational heterogeneity in the N-terminal domain of DGAT1, there is a small gain of secondary structure induced by ligand binding.

Authors:
 [1];  [2];  [1];  [1];  [1];  [1];  [1]
  1. Univ. of Alberta, Edmonton, AB (Canada)
  2. Stanford Univ., Menlo Park, CA (United States)
Publication Date:
Research Org.:
SLAC National Accelerator Lab., Menlo Park, CA (United States)
Sponsoring Org.:
USDOE
OSTI Identifier:
1490671
Grant/Contract Number:  
AC02-76SF00515
Resource Type:
Accepted Manuscript
Journal Name:
Scientific Reports
Additional Journal Information:
Journal Volume: 8; Journal Issue: 1; Journal ID: ISSN 2045-2322
Publisher:
Nature Publishing Group
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES

Citation Formats

Panigrahi, Rashmi, Matsui, Tsutomu, Song, Andrew H., Caldo, Kristian Mark P., Young, Howard S., Weselake, Randall J., and Lemieux, M. Joanne. Intrinsic disorder in the regulatory N-terminal domain of diacylglycerol acyltransferase 1 from Brassica napus. United States: N. p., 2018. Web. doi:10.1038/s41598-018-34339-1.
Panigrahi, Rashmi, Matsui, Tsutomu, Song, Andrew H., Caldo, Kristian Mark P., Young, Howard S., Weselake, Randall J., & Lemieux, M. Joanne. Intrinsic disorder in the regulatory N-terminal domain of diacylglycerol acyltransferase 1 from Brassica napus. United States. doi:10.1038/s41598-018-34339-1.
Panigrahi, Rashmi, Matsui, Tsutomu, Song, Andrew H., Caldo, Kristian Mark P., Young, Howard S., Weselake, Randall J., and Lemieux, M. Joanne. Mon . "Intrinsic disorder in the regulatory N-terminal domain of diacylglycerol acyltransferase 1 from Brassica napus". United States. doi:10.1038/s41598-018-34339-1. https://www.osti.gov/servlets/purl/1490671.
@article{osti_1490671,
title = {Intrinsic disorder in the regulatory N-terminal domain of diacylglycerol acyltransferase 1 from Brassica napus},
author = {Panigrahi, Rashmi and Matsui, Tsutomu and Song, Andrew H. and Caldo, Kristian Mark P. and Young, Howard S. and Weselake, Randall J. and Lemieux, M. Joanne},
abstractNote = {Proteins with multifunctional regulatory domains often demonstrate structural plasticity or protein disorder, allowing the binding of multiple regulatory factors and post-translational modifications. While the importance of protein disorder is clear, it also poses a challenge for in vitro characterization. Here, we report protein intrinsic disorder in a plant molecular system, which despite its prevalence is less studied. We present a detailed biophysical characterization of the entire cytoplasmic N-terminal domain of Brassica napus diacylglycerol acyltransferase, (DGAT1), which includes an inhibitory module and allosteric binding sites. Our results demonstrate that the monomeric N-terminal domain can be stabilized for biophysical characterization and is largely intrinsically disordered in solution. This domain interacts with allosteric modulators of DGAT1, CoA and oleoyl-CoA, at micromolar concentrations. Furthermore, while solution scattering studies indicate conformational heterogeneity in the N-terminal domain of DGAT1, there is a small gain of secondary structure induced by ligand binding.},
doi = {10.1038/s41598-018-34339-1},
journal = {Scientific Reports},
number = 1,
volume = 8,
place = {United States},
year = {2018},
month = {11}
}

Journal Article:
Free Publicly Available Full Text
Publisher's Version of Record

Figures / Tables:

Figure 1 Figure 1: Disorder propensity representation for the full-length BnaDGAT11-501 sequence (isoform BnaC. DGAT1) using DISOPRED. Disorder threshold is indicated by white line distinguishing between order and disorder. a). Inset 1: Cartoon representation of domains in BnaDGAT1; inset 2: A cartoon model of BnaDGAT1 embedded within the lipid bilayer, with themore » membrane domain depicted in green and the cytosolic domain depicted by a blue dashed line.« less

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Works referenced in this record:

Evolutionary Rate Heterogeneity in Proteins with Long Disordered Regions
journal, July 2002

  • Brown, Celeste J.; Takayama, Sachiko; Campen, Andrew M.
  • Journal of Molecular Evolution, Vol. 55, Issue 1
  • DOI: 10.1007/s00239-001-2309-6

Juxtanodin is an intrinsically disordered F-actin-binding protein
journal, November 2012

  • Ruskamo, Salla; Chukhlieb, Maryna; Vahokoski, Juha
  • Scientific Reports, Vol. 2, Issue 1
  • DOI: 10.1038/srep00899

Purification and properties of recombinant Brassica napus diacylglycerol acyltransferase 1
journal, February 2015


Why are ?natively unfolded? proteins unstructured under physiologic conditions?
journal, January 2000


Sequence complexity of disordered protein
journal, January 2000


Regulation of Cell Division by Intrinsically Unstructured Proteins: Intrinsic Flexibility, Modularity, and Signaling Conduits
journal, July 2008

  • Galea, Charles A.; Wang, Yuefeng; Sivakolundu, Sivashankar G.
  • Biochemistry, Vol. 47, Issue 29
  • DOI: 10.1021/bi8006803

Increasing the flow of carbon into seed oil
journal, November 2009


PRIMUS: a Windows PC-based system for small-angle scattering data analysis
journal, September 2003

  • Konarev, Petr V.; Volkov, Vladimir V.; Sokolova, Anna V.
  • Journal of Applied Crystallography, Vol. 36, Issue 5, p. 1277-1282
  • DOI: 10.1107/S0021889803012779

Large-Scale Comparative Phosphoproteomics Identifies Conserved Phosphorylation Sites in Plants
journal, May 2010

  • Nakagami, Hirofumi; Sugiyama, Naoyuki; Mochida, Keiichi
  • Plant Physiology, Vol. 153, Issue 3
  • DOI: 10.1104/pp.110.157347

Protein–protein interaction through β-strand addition
journal, August 2006


Intrinsically disordered proteins in cellular signalling and regulation
journal, December 2014

  • Wright, Peter E.; Dyson, H. Jane
  • Nature Reviews Molecular Cell Biology, Vol. 16, Issue 1
  • DOI: 10.1038/nrm3920

Small-angle scattering studies of intrinsically disordered proteins and their complexes
journal, February 2017

  • Cordeiro, Tiago N.; Herranz-Trillo, Fátima; Urbanek, Annika
  • Current Opinion in Structural Biology, Vol. 42
  • DOI: 10.1016/j.sbi.2016.10.011

Linking folding and binding
journal, February 2009


ATSAS 2.1 – towards automated and web-supported small-angle scattering data analysis
journal, April 2007

  • Petoukhov, Maxim V.; Konarev, Peter V.; Kikhney, Alexey G.
  • Journal of Applied Crystallography, Vol. 40, Issue s1
  • DOI: 10.1107/S0021889807002853

SEDPHAT – A platform for global ITC analysis and global multi-method analysis of molecular interactions
journal, April 2015


Controllable Activation of Nanoscale Dynamics in a Disordered Protein Alters Binding Kinetics
journal, April 2017

  • Callaway, David J. E.; Matsui, Tsutomu; Weiss, Thomas
  • Journal of Molecular Biology, Vol. 429, Issue 7
  • DOI: 10.1016/j.jmb.2017.03.003

FoldIndex(C): a simple tool to predict whether a given protein sequence is intrinsically unfolded
journal, June 2005


FoXS, FoXSDock and MultiFoXS: Single-state and multi-state structural modeling of proteins and their complexes based on SAXS profiles
journal, May 2016

  • Schneidman-Duhovny, Dina; Hammel, Michal; Tainer, John A.
  • Nucleic Acids Research, Vol. 44, Issue W1
  • DOI: 10.1093/nar/gkw389

DISOPRED3: precise disordered region predictions with annotated protein-binding activity
journal, November 2014


Classification of Intrinsically Disordered Regions and Proteins
journal, December 2013

  • van der Lee, Robin; Buljan, Marija; Lang, Benjamin
  • Chemical Reviews, Vol. 114, Issue 13
  • DOI: 10.1021/cr400525m

Understanding eukaryotic linear motifs and their role in cell signaling and regulation
journal, January 2008

  • Diella, Francesca
  • Frontiers in Bioscience, Vol. Volume, Issue 13
  • DOI: 10.2741/3175

Analysis of Molecular Recognition Features (MoRFs)
journal, October 2006

  • Mohan, Amrita; Oldfield, Christopher J.; Radivojac, Predrag
  • Journal of Molecular Biology, Vol. 362, Issue 5
  • DOI: 10.1016/j.jmb.2006.07.087

Modulation of Intrinsically Disordered Protein Function by Post-translational Modifications
journal, February 2016

  • Bah, Alaji; Forman-Kay, Julie D.
  • Journal of Biological Chemistry, Vol. 291, Issue 13
  • DOI: 10.1074/jbc.R115.695056

Understanding protein non-folding
journal, June 2010

  • Uversky, Vladimir N.; Dunker, A. Keith
  • Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics, Vol. 1804, Issue 6
  • DOI: 10.1016/j.bbapap.2010.01.017

Trifluoroethanol-induced conformational transitions of proteins: Insights gained from the differences between α-lactalbumin and ribonuclease A
journal, January 1999

  • Gast, Klaus; Zirwer, Dietrich; Müller-Frohne, Marlies
  • Protein Science, Vol. 8, Issue 3
  • DOI: 10.1110/ps.8.3.625

MultIDIMensionality of IDIMs: Intrinsic Disorder in Autoinhibition
journal, March 2013


A Bimodal Distribution of Two Distinct Categories of Intrinsically Disordered Structures with Separate Functions in FG Nucleoporins
journal, April 2010

  • Yamada, Justin; Phillips, Joshua L.; Patel, Samir
  • Molecular & Cellular Proteomics, Vol. 9, Issue 10
  • DOI: 10.1074/mcp.M000035-MCP201

Direct measurement of protein binding energetics by isothermal titration calorimetry
journal, September 2001


Estimation of Protein Secondary Structure from Circular Dichroism Spectra: Comparison of CONTIN, SELCON, and CDSSTR Methods with an Expanded Reference Set
journal, December 2000

  • Sreerama, Narasimha; Woody, Robert W.
  • Analytical Biochemistry, Vol. 287, Issue 2, p. 252-260
  • DOI: 10.1006/abio.2000.4880

Structural Basis of the pH-Dependent Assembly of a Botulinum Neurotoxin Complex
journal, November 2014


A decade and a half of protein intrinsic disorder: Biology still waits for physics: Protein Intrinsic Disorder
journal, April 2013


Malleable machines take shape in eukaryotic transcriptional regulation
journal, November 2008

  • Fuxreiter, Monika; Tompa, Peter; Simon, István
  • Nature Chemical Biology, Vol. 4, Issue 12
  • DOI: 10.1038/nchembio.127

Intrinsically Disordered Regions of Human Plasma Membrane Proteins Preferentially Occur in the Cytoplasmic Segment
journal, May 2007

  • Minezaki, Yoshiaki; Homma, Keiichi; Nishikawa, Ken
  • Journal of Molecular Biology, Vol. 368, Issue 3
  • DOI: 10.1016/j.jmb.2007.02.033

Protein secondary structure analyses from circular dichroism spectroscopy: Methods and reference databases
journal, May 2008

  • Whitmore, Lee; Wallace, B. A.
  • Biopolymers, Vol. 89, Issue 5, p. 392-400
  • DOI: 10.1002/bip.20853

Diacylglycerol Acyltransferase 1 Is Regulated by Its N-Terminal Domain in Response to Allosteric Effectors
journal, August 2017

  • Caldo, Kristian Mark P.; Acedo, Jeella Z.; Panigrahi, Rashmi
  • Plant Physiology, Vol. 175, Issue 2
  • DOI: 10.1104/pp.17.00934

An N-terminal fragment of mouse DGAT1 binds different acyl-CoAs with varying affinity
journal, August 2008

  • Siloto, Rodrigo M. P.; Madhavji, Milan; Wiehler, William B.
  • Biochemical and Biophysical Research Communications, Vol. 373, Issue 3
  • DOI: 10.1016/j.bbrc.2008.06.031

Analysis of structured and intrinsically disordered regions of transmembrane proteins
journal, January 2009

  • Xue, Bin; Li, Liwei; Meroueh, Samy O.
  • Molecular BioSystems, Vol. 5, Issue 12
  • DOI: 10.1039/b905913j

Intrinsically unstructured proteins
journal, October 2002


Protein–ligand interactions investigated by thermal shift assays (TSA) and dual polarization interferometry (DPI)
journal, January 2015

  • Grøftehauge, Morten K.; Hajizadeh, Nelly R.; Swann, Marcus J.
  • Acta Crystallographica Section D Biological Crystallography, Vol. 71, Issue 1
  • DOI: 10.1107/s1399004714016617

Intrinsic Disorder in Transmembrane Proteins: Roles in Signaling and Topology Prediction
journal, July 2016


Lipid binding by the Unique and SH3 domains of c-Src suggests a new regulatory mechanism
journal, February 2013

  • Pérez, Yolanda; Maffei, Mariano; Igea, Ana
  • Scientific Reports, Vol. 3, Issue 1
  • DOI: 10.1038/srep01295

GlobPlot: exploring protein sequences for globularity and disorder
journal, July 2003


Prediction and Functional Analysis of Native Disorder in Proteins from the Three Kingdoms of Life
journal, March 2004


Protein Posttranslational Modifications: The Chemistry of Proteome Diversifications
journal, November 2005

  • Walsh, Christopher T.; Garneau-Tsodikova, Sylvie; Gatto, Gregory J.
  • Angewandte Chemie International Edition, Vol. 44, Issue 45
  • DOI: 10.1002/anie.200501023

Folding patterns of porin and bacteriorhodopsin.
journal, June 1985


Structural analysis of intrinsically disordered proteins by small-angle X-ray scattering
journal, January 2012

  • Bernadó, Pau; Svergun, Dmitri I.
  • Mol. BioSyst., Vol. 8, Issue 1
  • DOI: 10.1039/c1mb05275f

Protein intrinsic disorder in plants
journal, January 2013

  • Pazos, Florencio; Pietrosemoli, Natalia; García-Martín, Juan A.
  • Frontiers in Plant Science, Vol. 4
  • DOI: 10.3389/fpls.2013.00363

Metabolic control analysis is helpful for informed genetic manipulation of oilseed rape (Brassica napus) to increase seed oil content
journal, August 2008

  • Weselake, Randall J.; Shah, Saleh; Tang, Mingguo
  • Journal of Experimental Botany, Vol. 59, Issue 13
  • DOI: 10.1093/jxb/ern206

Diacylglycerol acyltransferase 1 is activated by phosphatidate and inhibited by SnRK1-catalyzed phosphorylation
journal, August 2018

  • Caldo, Kristian Mark P.; Shen, Wei; Xu, Yang
  • The Plant Journal, Vol. 96, Issue 2
  • DOI: 10.1111/tpj.14029

Intrinsically disordered proteins: regulation and disease
journal, June 2011

  • Babu, M. Madan; van der Lee, Robin; de Groot, Natalia Sanchez
  • Current Opinion in Structural Biology, Vol. 21, Issue 3
  • DOI: 10.1016/j.sbi.2011.03.011

MoRFchibi SYSTEM: software tools for the identification of MoRFs in protein sequences
journal, May 2016

  • Malhis, Nawar; Jacobson, Matthew; Gsponer, Jörg
  • Nucleic Acids Research, Vol. 44, Issue W1
  • DOI: 10.1093/nar/gkw409

Sequence Determinants of Compaction in Intrinsically Disordered Proteins
journal, May 2010


Mechanism of One-to-Many Molecular Recognition Accompanying Target-Dependent Structure Formation: For the Tumor Suppressor p53 Protein as an Example
journal, October 2015

  • Hayashi, Tomohiko; Oshima, Hiraku; Yasuda, Satoshi
  • The Journal of Physical Chemistry B, Vol. 119, Issue 44
  • DOI: 10.1021/acs.jpcb.5b08513

GPS-Lipid: a robust tool for the prediction of multiple lipid modification sites
journal, June 2016

  • Xie, Yubin; Zheng, Yueyuan; Li, Hongyu
  • Scientific Reports, Vol. 6, Issue 1
  • DOI: 10.1038/srep28249

Analysis of Protein Stability and Ligand Interactions by Thermal Shift Assay
journal, February 2015


Engineering increased triacylglycerol accumulation in Saccharomyces cerevisiae using a modified type 1 plant diacylglycerol acyltransferase
journal, December 2014

  • Greer, Michael S.; Truksa, Martin; Deng, Wei
  • Applied Microbiology and Biotechnology, Vol. 99, Issue 5
  • DOI: 10.1007/s00253-014-6284-4

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