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Title: Rational Design Principles for the Transport and Subcellular Distribution of Nanomaterials into Plant Protoplasts

Abstract

Abstract The ability to control the subcellular localization of nanoparticles within living plants offers unique advantages for targeted biomolecule delivery and enables important applications in plant bioengineering. However, the mechanism of nanoparticle transport past plant biological membranes is poorly understood. Here, a mechanistic study of nanoparticle cellular uptake into plant protoplasts is presented. An experimentally validated mathematical model of lipid exchange envelope penetration mechanism for protoplasts, which predicts that the subcellular distribution of nanoparticles in plant cells is dictated by the particle size and the magnitude of the zeta potential, is advanced. The mechanism is completely generic, describing nanoparticles ranging from quantum dots, gold and silica nanoparticles, nanoceria, and single‐walled carbon nanotubes (SWNTs). In addition, the use of imaging flow cytometry to investigate the influence of protoplasts' morphological characteristics on nanoparticle uptake efficiency is demonstrated. Using DNA‐wrapped SWNTs as model nanoparticles, it is found that glycerolipids, the predominant lipids in chloroplast membranes, exhibit stronger lipid–nanoparticle interaction than phospholipids, the major constituent in protoplast membrane. This work can guide the rational design of nanoparticles for targeted delivery into specific compartments within plant cells without the use of chemical or mechanical aid, potentially enabling various plant engineering applications.

Authors:
 [1];  [1];  [1];  [1];  [1];  [1]
  1. Massachusetts Inst. of Technology (MIT), Cambridge, MA (United States)
Publication Date:
Research Org.:
Massachusetts Inst. of Technology (MIT), Cambridge, MA (United States)
Sponsoring Org.:
USDOE Office of Science (SC)
OSTI Identifier:
1609981
Alternate Identifier(s):
OSTI ID: 1469026
Grant/Contract Number:  
FG02-08ER46488; DE‐FG02‐08ER46488 Mod 0008
Resource Type:
Accepted Manuscript
Journal Name:
Small
Additional Journal Information:
Journal Volume: 14; Journal Issue: 44; Journal ID: ISSN 1613-6810
Publisher:
Wiley
Country of Publication:
United States
Language:
English
Subject:
36 MATERIALS SCIENCE; Chemistry; Science & Technology; Materials Science; Physics

Citation Formats

Lew, Tedrick Thomas Salim, Wong, Min Hao, Kwak, Seon-Yeong, Sinclair, Rosalie, Koman, Volodymyr B., and Strano, Michael S. Rational Design Principles for the Transport and Subcellular Distribution of Nanomaterials into Plant Protoplasts. United States: N. p., 2018. Web. doi:10.1002/smll.201802086.
Lew, Tedrick Thomas Salim, Wong, Min Hao, Kwak, Seon-Yeong, Sinclair, Rosalie, Koman, Volodymyr B., & Strano, Michael S. Rational Design Principles for the Transport and Subcellular Distribution of Nanomaterials into Plant Protoplasts. United States. https://doi.org/10.1002/smll.201802086
Lew, Tedrick Thomas Salim, Wong, Min Hao, Kwak, Seon-Yeong, Sinclair, Rosalie, Koman, Volodymyr B., and Strano, Michael S. Thu . "Rational Design Principles for the Transport and Subcellular Distribution of Nanomaterials into Plant Protoplasts". United States. https://doi.org/10.1002/smll.201802086. https://www.osti.gov/servlets/purl/1609981.
@article{osti_1609981,
title = {Rational Design Principles for the Transport and Subcellular Distribution of Nanomaterials into Plant Protoplasts},
author = {Lew, Tedrick Thomas Salim and Wong, Min Hao and Kwak, Seon-Yeong and Sinclair, Rosalie and Koman, Volodymyr B. and Strano, Michael S.},
abstractNote = {Abstract The ability to control the subcellular localization of nanoparticles within living plants offers unique advantages for targeted biomolecule delivery and enables important applications in plant bioengineering. However, the mechanism of nanoparticle transport past plant biological membranes is poorly understood. Here, a mechanistic study of nanoparticle cellular uptake into plant protoplasts is presented. An experimentally validated mathematical model of lipid exchange envelope penetration mechanism for protoplasts, which predicts that the subcellular distribution of nanoparticles in plant cells is dictated by the particle size and the magnitude of the zeta potential, is advanced. The mechanism is completely generic, describing nanoparticles ranging from quantum dots, gold and silica nanoparticles, nanoceria, and single‐walled carbon nanotubes (SWNTs). In addition, the use of imaging flow cytometry to investigate the influence of protoplasts' morphological characteristics on nanoparticle uptake efficiency is demonstrated. Using DNA‐wrapped SWNTs as model nanoparticles, it is found that glycerolipids, the predominant lipids in chloroplast membranes, exhibit stronger lipid–nanoparticle interaction than phospholipids, the major constituent in protoplast membrane. This work can guide the rational design of nanoparticles for targeted delivery into specific compartments within plant cells without the use of chemical or mechanical aid, potentially enabling various plant engineering applications.},
doi = {10.1002/smll.201802086},
journal = {Small},
number = 44,
volume = 14,
place = {United States},
year = {Thu Sep 06 00:00:00 EDT 2018},
month = {Thu Sep 06 00:00:00 EDT 2018}
}

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H.; Baulin, Vladimir A.</span> </li> <li> arXiv</li> <li> <span class="text-muted related-url">DOI: <a href="https://doi.org/10.48550/arxiv.1202.2314" class="text-muted" target="_blank" rel="noopener noreferrer">10.48550/arxiv.1202.2314<span class="fa fa-external-link" aria-hidden="true"></span></a></span> </li> </ul> <hr/> </div> </div> <div class="pagination-container small"> <a class="pure-button prev page" href="#" rel="prev"><span class="sr-only">Previous Page</span><span class="fa fa-angle-left"></span></a> <ul class="pagination d-inline-block" style="padding-left:.2em;"></ul> <a class="pure-button next page" href="#" rel="next"><span class="sr-only">Next Page</span><span class="fa fa-angle-right"></span></a> </div> </div> </div> <div class="col-sm-3 order-sm-3"> <ul class="nav nav-stacked"> <li class="active"><a href="" class="reference-type-filter tab-nav" data-tab="biblio-references" data-filter="type" data-pattern="*"><span class="fa fa-angle-right"></span> All References</a></li> <li class="small" style="margin-left:.75em; 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font-size:0.75rem;"><br/> <span class="type">journal</span>, <span class="date" data-date="2019-11-08">November 2019</span></small> </h2> <ul class="small references-list" style="list-style-type:none; margin-top: 0.5em; padding-left: 0; line-height:1.8em;"> <li> <span style="color:#5C7B2D;"> Lew, Tedrick Thomas Salim; Koman, Volodymyr B.; Gordiichuk, Pavlo</span> </li> <li> Advanced Materials Technologies, Vol. 5, Issue 3</li> <li> <span class="text-muted related-url">DOI: <a href="https://doi.org/10.1002/admt.201900657" class="text-muted" target="_blank" rel="noopener noreferrer">10.1002/admt.201900657<span class="fa fa-external-link" aria-hidden="true"></span></a></span> </li> </ul> <hr/> </div> <div> <h2 class="title" style="margin-bottom:0;" data-apporder=""> <a href="https://doi.org/10.1007/s12274-019-2438-0" target="_blank" rel="noopener noreferrer" class="name">Enhancing bioelectricity generation in microbial fuel cells and biophotovoltaics using nanomaterials<span class="fa fa-external-link" aria-hidden="true"></span></a> <small class="text-muted" style="text-transform:uppercase; font-size:0.75rem;"><br/> <span class="type">journal</span>, <span class="date" data-date="2019-06-11">June 2019</span></small> </h2> <ul class="small references-list" style="list-style-type:none; margin-top: 0.5em; padding-left: 0; line-height:1.8em;"> <li> <span style="color:#5C7B2D;"> Mouhib, Mohammed; Antonucci, Alessandra; Reggente, Melania</span> </li> <li> Nano Research, Vol. 12, Issue 9</li> <li> <span class="text-muted related-url">DOI: <a href="https://doi.org/10.1007/s12274-019-2438-0" class="text-muted" target="_blank" rel="noopener noreferrer">10.1007/s12274-019-2438-0<span class="fa fa-external-link" aria-hidden="true"></span></a></span> </li> </ul> <hr/> </div> <div> <h2 class="title" style="margin-bottom:0;" data-apporder=""> <a href="https://doi.org/10.1038/s41565-019-0375-4" target="_blank" rel="noopener noreferrer" class="name">Chloroplast-selective gene delivery and expression in planta using chitosan-complexed single-walled carbon nanotube carriers<span class="fa fa-external-link" aria-hidden="true"></span></a> <small class="text-muted" style="text-transform:uppercase; font-size:0.75rem;"><br/> <span class="type">journal</span>, <span class="date" data-date="2019-02-25">February 2019</span></small> </h2> <ul class="small references-list" style="list-style-type:none; margin-top: 0.5em; padding-left: 0; line-height:1.8em;"> <li> <span style="color:#5C7B2D;"> Kwak, Seon-Yeong; Lew, Tedrick Thomas Salim; Sweeney, Connor J.</span> </li> <li> Nature Nanotechnology, Vol. 14, Issue 5</li> <li> <span class="text-muted related-url">DOI: <a href="https://doi.org/10.1038/s41565-019-0375-4" class="text-muted" target="_blank" rel="noopener noreferrer">10.1038/s41565-019-0375-4<span class="fa fa-external-link" aria-hidden="true"></span></a></span> </li> </ul> <hr/> </div> <div> <h2 class="title" style="margin-bottom:0;" data-apporder=""> <a href="https://doi.org/10.1038/s41565-019-0382-5" target="_blank" rel="noopener noreferrer" class="name">High aspect ratio nanomaterials enable delivery of functional genetic material without DNA integration in mature plants<span class="fa fa-external-link" aria-hidden="true"></span></a> <small class="text-muted" style="text-transform:uppercase; font-size:0.75rem;"><br/> <span class="type">journal</span>, <span class="date" data-date="2019-02-25">February 2019</span></small> </h2> <ul class="small references-list" style="list-style-type:none; margin-top: 0.5em; padding-left: 0; line-height:1.8em;"> <li> <span style="color:#5C7B2D;"> Demirer, Gozde S.; Zhang, Huan; Matos, Juliana L.</span> </li> <li> Nature Nanotechnology, Vol. 14, Issue 5</li> <li> <span class="text-muted related-url">DOI: <a href="https://doi.org/10.1038/s41565-019-0382-5" class="text-muted" target="_blank" rel="noopener noreferrer">10.1038/s41565-019-0382-5<span class="fa fa-external-link" aria-hidden="true"></span></a></span> </li> </ul> <hr/> </div> </div> <div class="pagination-container small"> <a class="pure-button prev page" href="#" rel="prev"><span class="sr-only">Previous Page</span><span class="fa fa-angle-left"></span></a> <ul class="pagination d-inline-block" style="padding-left:.2em;"></ul> <a class="pure-button next page" href="#" rel="next"><span class="sr-only">Next Page</span><span class="fa fa-angle-right"></span></a> </div> </div> </div> <div class="col-sm-3 order-sm-3"> <ul class="nav nav-stacked"> <li class="active"><a href="" class="reference-type-filter tab-nav" data-filter="type" data-pattern="*"><span class="fa fa-angle-right"></span> All Cited By</a></li> <li class="small" style="margin-left:.75em; 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float:none;">[ × clear filter / sort ]</a> </div> <input type="submit" id="sort_submit_citations" name="submit" aria-label="submit" style="display: none;"/> </form> </div> </div> </div> </section> <section id="biblio-related" class="tab-content tab-content-sec " data-tab="biblio"> <div class="row"> <div class="col-sm-9 order-sm-9"> <section id="biblio-similar" class="tab-content tab-content-sec active" data-tab="related"> <div class="padding"> <p class="lead text-muted" style="font-size: 18px; margin-top:0px;">Similar Records in DOE PAGES and OSTI.GOV collections:</p> <aside> <ul class="item-list" itemscope itemtype="http://schema.org/ItemList" style="padding-left:0; list-style-type: none;"> <li> <div class="article item document" itemprop="itemListElement" itemscope itemtype="http://schema.org/WebPage"><meta itemprop="position" content="1" /><div class="item-info"> <h2 class="title" itemprop="name headline"><a href="/pages/biblio/1506633-effect-ceo2-nanomaterial-surface-functional-groups-tissue-subcellular-distribution-ce-tomato-solanum-lycopersicum" itemprop="url">Effect of CeO<sub>2</sub> nanomaterial surface functional groups on tissue and subcellular distribution of Ce in tomato (<em>Solanum lycopersicum</em>)</a></h2> <div class="metadata"> <small class="text-muted" style="text-transform:uppercase;display:block;line-height:2.5em;">Journal Article</small><span class="authors"> <span class="author">Li, Jieran</span> ; <span class="author">Tappero, Ryan V.</span> ; <span class="author">Acerbo, Alvin S.</span> ; <span class="author">...</span> <span class="text-muted pubdata"> - Environmental Science: Nano</span> </span> </div> <div class="abstract">Using recent advances in X-ray microscopy, this study aimed to elucidate mechanisms of uptake, subcellular distribution, and translocation of functionalized CeO<sub>2</sub> MNM (manufactured nanomaterials), having different charges, by tomato plants (<em>Solanum lycopersicum</em> cv Micro-Tom). As a result, we found that plant growth and Ce concentration in tissues were functions of surface charge and exposure concentration with root to shoot translocation being much greater for negatively charged CeO<sub>2</sub> than positive or neutral CeO<sub>2</sub>. Mechanisms of entry into roots and translocation within plants were examined using X-ray nano- and microprobes. There were dramatic differences in the tissue and subcellular distributions of Ce<a href='#' onclick='$(this).hide().next().show().next().show();return false;' style='margin-left:10px;'>more »</a><span style='display:none;'> in plant roots exposed to dextran-coated CeO<sub>2</sub> nanoparticles conjugated with positive, neutral and negative functional groups. Positively charged CeO<sub>2</sub> remained mainly bound to the epidermis of the root with little present in the apoplast or cytoplasm. Negatively charged CeO<sub>2</sub> was found in the cytoplasm throughout the root cross section, and negatively charged CeO<sub>2</sub> was found within the apoplast in the cortex and both the apoplast and the cytoplasm in the vasculature. Neutral CeO<sub>2</sub> likely entered through the gaps between epidermal cells being sloughed off during root growth and penetrated deeper into the interior of the roots (vasculature) via a combination of apoplastic and symplastic transport. Evidence of symplastic Ce transport was observed with the neutrally and negatively charged particles. We observed evidence of endocytosis as the mechanism for entry into the symplast allowing for entry into the xylem. Finally, this study provides critical information on how particle surface chemistry influences the biodistribution and cellular localization of nanomaterials in plants and is to date the highest resolution X-ray imaging of nanomaterials in plant cells.</span><a href='#' onclick='$(this).hide().prev().hide().prev().show();return false;' style='margin-left:10px;display:none;'>« less</a></div><div class="metadata-links small clearfix text-muted" style="margin-top:15px;"> <span class="fa fa-book text-muted" aria-hidden="true"></span> Cited by 31<div class="pure-menu pure-menu-horizontal pull-right" style="width:unset;"> <ul class="pure-menu-list"> <li class="pure-menu-item"><span class="item-info-ftlink"><a class="misc doi-link " href="https://doi.org/10.1039/C8EN01287C" target="_blank" rel="noopener" title="Link to document DOI" data-ostiid="1506633" data-product-type="Journal Article" data-product-subtype="AM" >https://doi.org/10.1039/C8EN01287C</a></span></li> <li class="pure-menu-item"><span class="item-info-ftlink"><a class="misc fulltext-link " href="/pages/servlets/purl/1506633" title="Link to document media" target="_blank" rel="noopener" data-ostiid="1506633" data-product-type="Journal Article" data-product-subtype="AM" >Full Text Available</a></span></li> </ul> </div> </div> </div> <div class="clearfix"></div> </div> </li> <li> <div class="article item document" itemprop="itemListElement" itemscope itemtype="http://schema.org/WebPage"><meta itemprop="position" content="2" /><div class="item-info"> <h2 class="title" itemprop="name headline"><a href="/pages/biblio/1836619-carbon-nanotube-biocompatibility-plants-determined-surface-chemistry" itemprop="url">Carbon nanotube biocompatibility in plants is determined by their surface chemistry</a></h2> <div class="metadata"> <small class="text-muted" style="text-transform:uppercase;display:block;line-height:2.5em;">Journal Article</small><span class="authors"> <span class="author">González-Grandío, Eduardo</span> ; <span class="author">Demirer, Gözde S.</span> ; <span class="author">Jackson, Christopher T.</span> ; <span class="author">...</span> <span class="text-muted pubdata"> - Journal of Nanobiotechnology</span> </span> </div> <div class="abstract">Abstract Background Agriculture faces significant global challenges including climate change and an increasing food demand due to a growing population. Addressing these challenges will require the adoption of transformative innovations into biotechnology practice, such as nanotechnology. Recently, nanomaterials have emerged as unmatched tools for their use as biosensors, or as biomolecule delivery vehicles. Despite their increasingly prolific use, plant-nanomaterial interactions remain poorly characterized, drawing into question the breadth of their utility and their broader environmental compatibility. Results Herein, we characterize the response of Arabidopsis thaliana to single walled carbon nanotube (SWNT) exposure with two different surface chemistries commonly used for biosensing and<a href='#' onclick='$(this).hide().next().show().next().show();return false;' style='margin-left:10px;'>more »</a><span style='display:none;'> nucleic acid delivery: oligonucleotide adsorbed-pristine SWNTs, and polyethyleneimine-SWNTs loaded with plasmid DNA (PEI-SWNTs), both introduced by leaf infiltration. We observed that pristine SWNTs elicit a mild stress response almost undistinguishable from the infiltration process, indicating that these nanomaterials are well-tolerated by the plant. However, PEI-SWNTs induce a much larger transcriptional reprogramming that involves stress, immunity, and senescence responses. PEI-SWNT-induced transcriptional profile is very similar to that of mutant plants displaying a constitutive immune response or treated with stress-priming agrochemicals. We selected molecular markers from our transcriptomic analysis and identified PEI as the main cause of this adverse reaction. We show that PEI-SWNT response is concentration-dependent and, when persistent over time, leads to cell death. We probed a panel of PEI variant-functionalized SWNTs across two plant species and identified biocompatible SWNT surface functionalizations. Conclusions While SWNTs themselves are well tolerated by plants, SWNTs surface-functionalized with positively charged polymers become toxic and produce cell death. We use molecular markers to identify more biocompatible SWNT formulations. Our results highlight the importance of nanoparticle surface chemistry on their biocompatibility and will facilitate the use of functionalized nanomaterials for agricultural improvement. Graphical Abstract</span><a href='#' onclick='$(this).hide().prev().hide().prev().show();return false;' style='margin-left:10px;display:none;'>« less</a></div><div class="metadata-links small clearfix text-muted" style="margin-top:15px;"> <div class="pure-menu pure-menu-horizontal pull-right" style="width:unset;"> <ul class="pure-menu-list"> <li class="pure-menu-item"><span class="item-info-ftlink"><a class="misc doi-link " href="https://doi.org/10.1186/s12951-021-01178-8" target="_blank" rel="noopener" title="Link to document DOI" data-ostiid="1836619" data-product-type="Journal Article" data-product-subtype="PA" >https://doi.org/10.1186/s12951-021-01178-8</a></span></li> </ul> </div> </div> </div> <div class="clearfix"></div> </div> </li> <li> <div class="article item document" itemprop="itemListElement" itemscope itemtype="http://schema.org/WebPage"><meta itemprop="position" content="3" /><div class="item-info"> <h2 class="title" itemprop="name headline"><a href="/pages/biblio/1643174-lights-up-organelles-optogenetic-tools-control-subcellular-structure-organization" itemprop="url">Lights up on organelles: Optogenetic tools to control subcellular structure and organization</a></h2> <div class="metadata"> <small class="text-muted" style="text-transform:uppercase;display:block;line-height:2.5em;">Journal Article</small><span class="authors"> <span class="author">Kichuk, Therese C.</span> ; <span class="author">Carrasco‐López, César</span> ; <span class="author">Avalos, José L.</span> <span class="text-muted pubdata"> - WIREs Mechanisms of Disease</span> </span> </div> <div class="abstract">Abstract Since the neurobiological inception of optogenetics, light‐controlled molecular perturbations have been applied in many scientific disciplines to both manipulate and observe cellular function. Proteins exhibiting light‐sensitive conformational changes provide researchers with avenues for spatiotemporal control over the cellular environment and serve as valuable alternatives to chemically inducible systems. Optogenetic approaches have been developed to target proteins to specific subcellular compartments, allowing for the manipulation of nuclear translocation and plasma membrane morphology. Additionally, these tools have been harnessed for molecular interrogation of organelle function, location, and dynamics. Optogenetic approaches offer novel ways to answer fundamental biological questions and to improve<a href='#' onclick='$(this).hide().next().show().next().show();return false;' style='margin-left:10px;'>more »</a><span style='display:none;'> the efficiency of bioengineered cell factories by controlling the assembly of synthetic organelles. This review first provides a summary of available optogenetic systems with an emphasis on their organelle‐specific utility. It then explores the strategies employed for organelle targeting and concludes by discussing our perspective on the future of optogenetics to control subcellular structure and organization. This article is categorized under: Metabolic Diseases > Molecular and Cellular Physiology</span><a href='#' onclick='$(this).hide().prev().hide().prev().show();return false;' style='margin-left:10px;display:none;'>« less</a></div><div class="metadata-links small clearfix text-muted" style="margin-top:15px;"> <span class="fa fa-book text-muted" aria-hidden="true"></span> Cited by 13<div class="pure-menu pure-menu-horizontal pull-right" style="width:unset;"> <ul class="pure-menu-list"> <li class="pure-menu-item"><span class="item-info-ftlink"><a class="misc doi-link " href="https://doi.org/10.1002/wsbm.1500" target="_blank" rel="noopener" title="Link to document DOI" data-ostiid="1643174" data-product-type="Journal Article" data-product-subtype="PM" >https://doi.org/10.1002/wsbm.1500</a></span></li> </ul> </div> </div> </div> <div class="clearfix"></div> </div> </li> <li> <div class="article item document" itemprop="itemListElement" itemscope itemtype="http://schema.org/WebPage"><meta itemprop="position" content="4" /><div class="item-info"> <h2 class="title" itemprop="name headline"><a href="/biblio/5840280-effects-freezing-cold-acclimation-plasma-membrane-isolated-protoplasts" itemprop="url">Effects of freezing and cold acclimation on the plasma membrane of isolated protoplasts</a></h2> <div class="metadata"> <small class="text-muted" style="text-transform:uppercase;display:block;line-height:2.5em;">Technical Report</small><span class="authors"> <span class="author">Steponkus, P L</span> <span class="text-muted pubdata"></span> </span> </div> <div class="abstract">The principal goal of our program is to provide a mechanistic understanding of the cellular and molecular aspects of freezing injury and cold acclimation from a perspective of the structural and functional integrity of the plasma membrane -- the primary site of freezing injury in winter cereals. Our immediate goals are (1) to provide an understanding of the mechanism by which freeze-induced dehydration affects the formation of aparticulate domains and lamellar-to-hexagonal{sub {parallel}} phase transitions in the plasma membrane of NA protoplasts, (2) to characterize the cellular and molecular mechanisms by which cold acclimation and cryoprotectants preclude or diminish these alterations<a href='#' onclick='$(this).hide().next().show().next().show();return false;' style='margin-left:10px;'>more »</a><span style='display:none;'> in the plasma membrane of ACC protoplasts and (3) to elucidate the molecular basis for the lesion that limits the maximum freezing tolerance of cold-acclimated winter rye and which is believed to be the formation of domains of interdigitated lipids in the L{sub {beta}} phase. This past year our efforts have included (a) characterization of the ultrastructural changes in the plasma membrane that are associated with freezing injury of protoplasts isolated from cold-acclimated rye leaves; (b) determinations of the hydration characteristics of plasma membrane lipids and model lipid mixtures, including the thermal dependence of the hydration characteristics; (c) studies of dehydration-induced phase transitions and demixing in model systems of plasma membrane lipids; (d) differential scanning calorimetry studies to determine the amount of freezable/unfreezable water that is associated with lipids; and (e) preliminary cryo-SEM observations of in situ ice formation in rye leaves. 11 refs.</span><a href='#' onclick='$(this).hide().prev().hide().prev().show();return false;' style='margin-left:10px;display:none;'>« less</a></div><div class="metadata-links small clearfix text-muted" style="margin-top:15px;"> <div class="pure-menu pure-menu-horizontal pull-right" style="width:unset;"> </div> </div> </div> <div class="clearfix"></div> </div> </li> <li> <div class="article item document" itemprop="itemListElement" itemscope itemtype="http://schema.org/WebPage"><meta itemprop="position" content="5" /><div class="item-info"> <h2 class="title" itemprop="name headline"><a href="/pages/biblio/1987891-engineering-characterization-carbohydratebinding-modules-imaging-cellulose-fibrils-biosynthesis-plant-protoplasts" itemprop="url">Engineering and characterization of carbohydrate‐binding modules for imaging cellulose fibrils biosynthesis in plant protoplasts</a></h2> <div class="metadata"> <small class="text-muted" style="text-transform:uppercase;display:block;line-height:2.5em;">Journal Article</small><span class="authors"> <span class="author">Jayachandran, Dharanidaran</span> ; <span class="author">Smith, Peter</span> ; <span class="author">Irfan, Mohammad</span> ; <span class="author">...</span> <span class="text-muted pubdata"> - Biotechnology and Bioengineering</span> </span> </div> <div class="abstract">Abstract Carbohydrate binding modules (CBMs) are noncatalytic domains that assist tethered catalytic domains in substrate targeting. CBMs have therefore been used to visualize distinct polysaccharides present in the cell wall of plant cells and tissues. However, most previous studies provide a qualitative analysis of CBM‐polysaccharide interactions, with limited characterization of engineered tandem CBM designs for recognizing polysaccharides like cellulose and limited application of CBM‐based probes to visualize cellulose fibrils synthesis in model plant protoplasts with regenerating cell walls. Here, we examine the dynamic interactions of engineered type‐A CBMs from families 3a and 64 with crystalline cellulose‐I and phosphoric acid swollen<a href='#' onclick='$(this).hide().next().show().next().show();return false;' style='margin-left:10px;'>more »</a><span style='display:none;'> cellulose. We generated tandem CBM designs to determine various characteristic properties including binding reversibility toward cellulose‐I using equilibrium binding assays. To compute the adsorption ( nk <sub>on</sub> ) and desorption ( k <sub>off</sub> ) rate constants of single versus tandem CBM designs toward nanocrystalline cellulose, we employed dynamic kinetic binding assays using quartz crystal microbalance with dissipation. Our results indicate that tandem CBM3a exhibited the highest adsorption rate to cellulose and displayed reversible binding to both crystalline/amorphous cellulose, unlike other CBM designs, making tandem CBM3a better suited for live plant cell wall biosynthesis imaging applications. We used several engineered CBMs to visualize Arabidopsis thaliana protoplasts with regenerated cell walls using confocal laser scanning microscopy and wide‐field fluorescence microscopy. Lastly, we also demonstrated how CBMs as probe reagents can enable in situ visualization of cellulose fibrils during cell wall regeneration in Arabidopsis protoplasts.</span><a href='#' onclick='$(this).hide().prev().hide().prev().show();return false;' style='margin-left:10px;display:none;'>« less</a></div><div class="metadata-links small clearfix text-muted" style="margin-top:15px;"> <div class="pure-menu pure-menu-horizontal pull-right" style="width:unset;"> <ul class="pure-menu-list"> <li class="pure-menu-item"><span class="item-info-ftlink"><a class="misc doi-link " href="https://doi.org/10.1002/bit.28484" target="_blank" rel="noopener" title="Link to document DOI" data-ostiid="1987891" data-product-type="Journal Article" data-product-subtype="PA" >https://doi.org/10.1002/bit.28484</a></span></li> </ul> </div> </div> </div> <div class="clearfix"></div> </div> </li> </ul> </aside> </div> </section> </div> <div class="col-sm-3 order-sm-3"> <ul class="nav nav-stacked"> <li class="active"><a class="tab-nav disabled" data-tab="related" style="color: #636c72 !important; opacity: 1;"><span class="fa fa-angle-right"></span> Similar Records</a></li> </ul> </div> </div> </section> </div></div> </div> </div> </section> <footer class="" style="background-color:#f9f9f9;"> <div class="footer-minor"> <div class="container"> <hr class="footer-separator"/> <br/> <div class="col text-center mt-3"> <div class="pure-menu pure-menu-horizontal"> <ul class="pure-menu-list" id="footer-org-menu"> <li class="pure-menu-item"> <a href="https://energy.gov" target="_blank" rel="noopener noreferrer"> <img src="data:image/gif;base64,R0lGODlhAQABAIAAAP///wAAACH5BAEAAAAALAAAAAABAAEAAAICRAEAOw==" class="sprite sprite-footer-us-doe-min" alt="U.S. Department of Energy" /> </a> </li> <li class="pure-menu-item"> <a href="https://www.energy.gov/science/office-science" target="_blank" rel="noopener noreferrer"> <img src="data:image/gif;base64,R0lGODlhAQABAIAAAP///wAAACH5BAEAAAAALAAAAAABAAEAAAICRAEAOw==" class="sprite sprite-footer-office-of-science-min" alt="Office of Science" /> </a> </li> <li class="pure-menu-item"> <a href="https://www.osti.gov" target="_blank" rel="noopener noreferrer"> <img src="data:image/gif;base64,R0lGODlhAQABAIAAAP///wAAACH5BAEAAAAALAAAAAABAAEAAAICRAEAOw==" class="sprite sprite-footer-osti-min" alt="Office of Scientific and Technical Information" /> </a> </li> </ul> </div> </div> <div class="col text-center small" style="margin-top: 0.5em;margin-bottom:2.0rem;"> <div class="row justify-content-center" style="color:white"> <div class="pure-menu pure-menu-horizontal" style='white-space:normal'> <ul class="pure-menu-list"> <li class="pure-menu-item"><a href="https://www.osti.gov/disclaim" class="pure-menu-link" target="_blank" ref="noopener noreferrer"><span class="fa fa-institution"></span> Website Policies <span class="d-none d-sm-inline d-print-none" style="color:#737373;">/ Important Links</span></a></li> <li class="pure-menu-item" style='float:none;'><a href="/pages/contact" class="pure-menu-link"><span class="fa fa-comments-o"></span>Contact Us</a></li> <li class="d-block d-md-none mb-1"></li> <li class="pure-menu-item" style='float:none;'><a target="_blank" title="Vulnerability Disclosure Program" class="pure-menu-link" href="https://doe.responsibledisclosure.com/hc/en-us" rel="noopener noreferrer">Vulnerability Disclosure Program</a></li> <li class="d-block d-lg-none mb-1"></li> <li class="pure-menu-item" style="float:none;"><a href="https://www.facebook.com/ostigov" target="_blank" class="pure-menu-link social ext fa fa-facebook" rel="noopener noreferrer"><span class="sr-only" style="background-color: #fff; color: #333;">Facebook</span></a></li> <li class="pure-menu-item" style="float:none;"><a href="https://twitter.com/OSTIgov" target="_blank" class="pure-menu-link social ext fa fa-twitter" rel="noopener noreferrer"><span class="sr-only" style="background-color: #fff; color: #333;">Twitter</span></a></li> <li class="pure-menu-item" style="float:none;"><a href="https://www.youtube.com/user/ostigov" target="_blank" class="pure-menu-link social ext fa fa-youtube-play" rel="noopener noreferrer"><span class="sr-only" style="background-color: #fff; color: #333;">Youtube</span></a></li> </ul> </div> </div> </div> </div> </div> </footer> <link href="/pages/css/pages.fonts.240327.0205.css" rel="stylesheet"> <script src="/pages/js/pages.240327.0205.js"></script><noscript></noscript> <script defer src="/pages/js/pages.biblio.240327.0205.js"></script><noscript></noscript> <script defer src="/pages/js/lity.js"></script><noscript></noscript> <script async type="text/javascript" src="/pages/js/Universal-Federated-Analytics-Min.js?agency=DOE" id="_fed_an_ua_tag"></script><noscript></noscript> </body> <!-- DOE PAGES v.240327.0205 --> </html>