Purification, Characterization, and Structural Analysis of a Plant Low-Temperature-Induced Protein

Boothe, J. G.; Sönnichsen, Frank D.; Beus, Mitchel D. de; Johnson‐Flanagan, Anne M.

doi:10.1104/pp.113.2.367

Cited by 26 publications

(14 citation statements)

References 45 publications

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“…Other workers have observed that apparently unfolded LEA (or similar) proteins become more structured when water activity is decreased by e.g. trifluoroethanol (56,58,66), high salt concentration (54), or drying in the presence of sucrose (67). This has obvious physiological relevance in desiccation-tolerant systems, including the anhydrobiotic nematode A. avenae, but leads to the further question of what the functions of the folded, presumably partially dehydrated, LEA proteins might be.…”

Section: Discussionmentioning

confidence: 99%

Transition from Natively Unfolded to Folded State Induced by Desiccation in an Anhydrobiotic Nematode Protein

Goyal

Tisi

Basran

et al. 2003

Journal of Biological Chemistry

186

187

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Late embryogenesis abundant (LEA) proteins are associated with desiccation tolerance in resurrection plants and in plant seeds, and the recent discovery of a dehydration-induced Group 3 LEA-like gene in the nematode Aphelenchus avenae suggests a similar association in anhydrobiotic animals. Despite their importance, little is known about the structure of Group 3 LEA proteins, although computer modeling and secondary structure algorithms predict a largely ␣-helical monomer that forms coiled coil oligomers. We have therefore investigated the structure of the nematode protein, Aav-LEA1, in the first such analysis of a well characterized Group 3 LEA-like protein. Immunoblotting and subunit cross-linking experiments demonstrate limited oligomerization of AavLEA1, but analytical ultracentrifugation and gel filtration show that the vast majority of the protein is monomeric. Moreover, CD, fluorescence emission, and Fourier transform-infrared spectroscopy indicate an unstructured conformation for the nematode protein. Therefore, in solution, no evidence was found to support structure predictions; instead, Aav-LEA1 seems to be natively unfolded with a high degree of hydration and low compactness. Such proteins can, however, be induced to fold into more rigid structures by partner molecules or by altered physiological conditions. Because AavLEA1 is associated with desiccation stress, its Fourier transform-infrared spectrum in the dehydrated state was examined. A dramatic but reversible increase in ␣-helix and, possibly, coiled coil formation was observed on drying, indicating that computer predictions of secondary structure may be correct for the solid state. This unusual finding offers the possibility that structural shifts in Group 3 LEA proteins occur on dehydration, perhaps consistent with their role in anhydrobiosis.

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Section: Discussionmentioning

confidence: 99%

Transition from Natively Unfolded to Folded State Induced by Desiccation in an Anhydrobiotic Nematode Protein

Goyal

Tisi

Basran

et al. 2003

Journal of Biological Chemistry

186

187

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“…B, Levels of the B. napus BN28 protein in nonacclimated (NA) and cold-acclimated (CA) control and CBF-expressing transgenic B. napus plants. Total soluble protein (100 g) prepared from nonacclimated and 3-week coldacclimated plants was subjected to immunoblot analysis using antiserum raised to the BN28 polypeptide (Boothe et al, 1997). Numbers above each sample refer to the specific transgenic line tested.…”

Section: A Cbf Cold-response Pathway In Brassica Napusmentioning

confidence: 99%

“…Total soluble protein (100 g) was fractionated by 10% (w/v) acrylamide tricine SDS/ PAGE (Schägger and von Jagow, 1987) and transferred to 0.1-m nitrocellulose membranes by electroblotting (Towbin et al, 1979) as described (Artus et al, 1996). BN28 protein was detected using antiserum kindly provided by Anne Johnson (Boothe et al, 1997) and visualized using the enhanced chemiluminescence system (Amersham, Buckinghamshire, UK).…”

Section: Immunoblot Analysismentioning

confidence: 99%

Components of the Arabidopsis C-Repeat/Dehydration-Responsive Element Binding Factor Cold-Response Pathway Are Conserved inBrassica napus and Other Plant Species

Kleff²,

et al. 2001

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Many plants increase in freezing tolerance in response to low, nonfreezing temperatures, a phenomenon known as cold acclimation. Cold acclimation in Arabidopsis involves rapid cold-induced expression of the C-repeat/dehydrationresponsive element binding factor (CBF) transcriptional activators followed by expression of CBF-targeted genes that increase freezing tolerance. Here, we present evidence for a CBF cold-response pathway in Brassica napus. We show that B. napus encodes CBF-like genes and that transcripts for these genes accumulate rapidly in response to low temperature followed closely by expression of the cold-regulated Bn115 gene, an ortholog of the Arabidopsis CBF-targeted COR15a gene. Moreover, we show that constitutive overexpression of the Arabidopsis CBF genes in transgenic B. napus plants induces expression of orthologs of Arabidopsis CBF-targeted genes and increases the freezing tolerance of both nonacclimated and cold-acclimated plants. Transcripts encoding CBF-like proteins were also found to accumulate rapidly in response to low temperature in wheat (Triticum aestivum L. cv Norstar) and rye (Secale cereale L. cv Puma), which cold acclimate, as well as in tomato (Lycopersicon esculentum var. Bonny Best, Castle Mart, Micro-Tom, and D Huang), a freezing-sensitive plant that does not cold acclimate. An alignment of the CBF proteins from Arabidopsis, B. napus, wheat, rye, and tomato revealed the presence of conserved amino acid sequences, PKK/RPAGRxKFxETRHP and DSAWR, that bracket the AP2/EREBP DNA binding domains of the proteins and distinguish them from other members of the AP2/EREBP protein family. We conclude that components of the CBF cold-response pathway are highly conserved in flowering plants and not limited to those that cold acclimate.Plants vary greatly in their abilities to survive freezing temperatures (Sakai and Larcher, 1987). Whereas plants from tropical regions have essentially no capacity to withstand freezing, herbaceous plants from temperate regions can survive freezing at temperatures ranging from Ϫ5 to Ϫ30°C, depending on the species. It is significant that the maximum freezing tolerance of plants is not constitutive, but is induced in response to low temperatures (below approximately 10°C), a phenomenon known as "cold acclimation" (Hughes and Dunn, 1996;Thomashow, 1999). Nonacclimated wheat (Triticum aestivum L. cv Norstar) plants, for instance, are killed at freezing temperatures of about Ϫ5°C, but after cold acclimation, can survive temperatures down to about Ϫ20°C. Determining what accounts for the differences in freezing tolerance between plant species and the molecular basis of cold acclimation is of basic scientific interest and has the potential to provide new approaches to improve the freezing tolerance of plants, an important agronomic trait.A recent advance in understanding cold acclimation in Arabidopsis was the discovery of the C-repeat/dehydration-responsive element binding factor (CBF) cold-response pathway (see Thomashow, 2001). Arabidopsis encodes a small fami...

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“…At decreasing temperature, intensity of NMR signals also decreases leading to the identification of intracellular freezing in the plant tissues. In Brassica, NMR was used to analyze the structure of low temperature-induced protein (Boothe et al 1997). NMR spectroscopy quantifies liquid water.…”

Section: Temperaturementioning

confidence: 99%

Identification of Subcellular, Structural, and Metabolic Changes Through NMR

Dhar

Malviya

2015

Phenomics in Crop Plants: Trends, Options and Limitations

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Secondary metabolites are unique sources for flavors, nutraceuticals, pharmaceuticals, and industrial bioactive molecules which are biosynthesized in different plant tissues. These metabolites play a major role in the adaptation of plants to the prevailing environment, in overcoming stress conditions, and in defense to several unforeseen invasions. Identifying the biological components and their functions and multiple interactions between components to understand the cellular metabolism of a cell to meet its fluctuating demand for energy and materials has remained a challenging task. Based on traditional metabolic analysis, mapping of intracellular fluxes in metabolic networks is only possible with high-throughput techniques. Nuclear magnetic resonance (NMR) spectroscopy is a powerful and versatile tool that can provide information on the metabolites and their metabolic network. NMR has played a dominant role in the identification of an array of compounds from diverse environments and diverse ecogeographic domains. Plant biotic relationships which include host plant interaction and resistance for an eco-metabolomics have been developed with NMR approach. NMR can also be used to determine low-resolution structures of target-ligand complexes for natively unstructured proteins or membrane proteins that are not amenable to crystallographic approaches.

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Purification, Characterization, and Structural Analysis of a Plant Low-Temperature-Induced Protein

Cited by 26 publications

References 45 publications

Transition from Natively Unfolded to Folded State Induced by Desiccation in an Anhydrobiotic Nematode Protein

Transition from Natively Unfolded to Folded State Induced by Desiccation in an Anhydrobiotic Nematode Protein

Components of the Arabidopsis C-Repeat/Dehydration-Responsive Element Binding Factor Cold-Response Pathway Are Conserved inBrassica napus and Other Plant Species

Identification of Subcellular, Structural, and Metabolic Changes Through NMR

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