Molecular Cloning, Expression, and Functional Characterization of a Cystatin from Pineapple Stem

Shyu, Douglas J. H.; Chyan, Chia-Lin; Tzen, Jason T. C.; Chou, Wing‐Ming

doi:10.1271/bbb.68.1681

Cited by 23 publications

(12 citation statements)

References 38 publications

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“…At the lower concentrations (5 and 10 lg), the inhibitory activity of GST-AtCYSb was much stronger than that of GST-AtCYSa. These inhibitory activities were similar to those reported for other plant cystatins (Shyu et al 2004;Christova et al 2006). These results indicate that AtCYSa and AtCYSb encode Arabidopsis cystatins.…”

Section: Resultssupporting

confidence: 87%

Two cysteine proteinase inhibitors from Arabidopsis thaliana, AtCYSa and AtCYSb, increasing the salt, drought, oxidation and cold tolerance

2008

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Two cysteine proteinase inhibitors (cystatins) from Arabidopsis thaliana, designated AtCYSa and AtCYSb, were characterized. Recombinant GST-AtCYSa and GST-AtCYSb were expressed in Escherichia coli and purified. They inhibit the catalytic activity of papain, which is generally taken as evidence for cysteine proteinase inhibitor function. Northern blot analyses showed that the expressions of AtCYSa and AtCYSb gene in Arabidopsis cells and seedlings were strongly induced by multiple abiotic stresses from high salt, drought, oxidant, and cold. Interestingly, the promoter region of AtCYSa gene contains a dehydration-responsive element (DRE) and an abscisic acid (ABA)-responsive element (ABRE), which identifies it as a DREB1A and AREB target gene. Under normal conditions, AtCYSa was expressed in 35S: DREB1A and 35S: AREB1 plants at a higher level than in WT plants, while AtCYSa gene was expressed in 35S: DREB2A plants at the same level as in WT plants. Under stress conditions (salt, drought and cold), AtCYSa was expressed more in all three transgenic plants than in WT plants. Over-expression of AtCYSa and AtCYSb in transgenic yeast and Arabidopsis plants increased the resistance to high salt, drought, oxidative, and cold stresses. Taken together, these data raise the possibility of using AtCYSa and AtCYSb to genetically improve environmental stresses tolerance in plants.

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Section: Resultssupporting

confidence: 87%

Two cysteine proteinase inhibitors from Arabidopsis thaliana, AtCYSa and AtCYSb, increasing the salt, drought, oxidation and cold tolerance

2008

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“…Plant cysteine protease inhibitors or cystatins regulate endogenous cysteine protease activity during various biological processes including defence, response to biotic and abiotic stress, embryogenesis and seed germination, and in PCD (Shyu et al 2004). Generation of ROS plays a key role in PCD in both plant and animal cells; at low levels antioxidant enzymes are induced, though when ROS levels reach a critical threshold a PCD pathway is triggered.…”

Section: Ppd Down-regulated Genesmentioning

confidence: 99%

Towards identifying the full set of genes expressed during cassava post-harvest physiological deterioration

et al. 2007

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Storage roots of cassava (Manihot esculenta Crantz) exhibit a rapid post-harvest physiological deterioration (PPD) response that can occur within 24-72 h of harvest. PPD is an enzymatically mediated oxidative process with parallels to plant wound, senescence and defence responses. To characterise those genes that show significant change in expression during the PPD response we have used cDNA microarray technology to carry out a large-scale analysis of the cassava root transcriptome during the post-harvest period. We identified 72 non-redundant expressed sequence tags which showed altered regulation during the post-harvest period. Of these 63 were induced, whilst 9 were down-regulated. RNA blot analysis of selected genes was used to verify and extend the microarray data. Additional microarray hybridisation experiments allowed the identification of 21 root-specific and 24 root-wounding-specific sequences. Many of the up-regulated and PPD-specific expressed sequence tags were predicted to play a role in cellular processes including reactive oxygen species turnover, cell wall repair, programmed cell death, ion, water or metabolite transport, signal transduction or perception, stress response, metabolism and biosynthesis, and activation of protein synthesis.

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“…Here, cysteine in the extraction buffer is important to keep the activity of cysteine proteases because these proteases use a catalytic cysteine residue as a nucleophile during proteolysis and are activated by thiol compounds (20). Many reports on cysteine protease purification from pear, kiwifruit, or pineapple used various concentration of cysteine (1 to 100 mM) in the extraction buffer (11,21).…”

Section: Resultsmentioning

confidence: 99%

Functional characterization of purified pear protease and its proteolytic activities with casein and myofibrillar proteins

Nam

Kim

Walsh

et al. 2016

Food Sci Biotechnol

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This study was performed to characterize pear protease proteolytic activity and investigate the use of pear protease as a meat tenderizer. Pear protease was purified and stabilized by 5% dextrin during lyophilization (dry) or concentration (liquid). Pear protease was further characterized with respect to pH, thermodynamics, and enzyme kinetics. Pear protease was stable at a pH range of 5-8 with an optimum pH of 6.5. From Arrhenius plots, liquid protease showed higher temperature dependency (23.49 kJ/mol) than dry protease (18.62 kJ/mol) due to its higher activation energy. The kcat/Km, catalytic efficiency of enzyme, was similar with 2.9 and 2.7 µM/min with dry and liquid proteases. Pear protease was evaluated for its proteolytic activities with casein and beef myofibrillar proteins by individually and combination with fig and kiwifruit proteases. These result indicated that pear and kiwifruit proteases could be complementary to be a desirable product for meat tenderization.

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Molecular Cloning, Expression, and Functional Characterization of a Cystatin from Pineapple Stem

Cited by 23 publications

References 38 publications

Two cysteine proteinase inhibitors from Arabidopsis thaliana, AtCYSa and AtCYSb, increasing the salt, drought, oxidation and cold tolerance

Two cysteine proteinase inhibitors from Arabidopsis thaliana, AtCYSa and AtCYSb, increasing the salt, drought, oxidation and cold tolerance

Towards identifying the full set of genes expressed during cassava post-harvest physiological deterioration

Functional characterization of purified pear protease and its proteolytic activities with casein and myofibrillar proteins

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