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Identification and biochemical characterization of 20S proteasome in wheat roots under salt stress

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Abstract

In order to gain a better understanding of proteases state in wheat roots under salt stress, the roots of wheat (Triticum aestivum L., H6756) exposed to 300 mM NaCl were investigated at different days after treatment (DAT). The results showed that the content of soluble proteins decreased continuously, but the activity of total proteases increased gradually, and five root endopeptidase isoenzymes (RE1-5) were detected by natural gradient polyacrylamide gel electrophoresis (GPAGE) with supplement of gelatin as a substrate. Among five REs, RE1 was not only detected in salt-treated roots, but also obviously possessed the higher activity, whereas its protein amount decreased gradually during the salt treatment process. The biochemical characters of RE1 were examined afterwards. The results showed that its molecular mass was about 740 kDa and its optimal temperature was about 40°C. But the optimal pH was 5 to substrate gelatin or 7–8 to substrate casein. Its activity was obviously inhibited by 10 mM PMSF and 20 μM MG115, and RE1 was further confirmed as 20S proteasome by Western blotting. The results in this study suggested that 20S proteasome of wheat roots might be an important protease accumulated in roots in response to salt stress.

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Abbreviations

DAT:

days after treatment

GPAGE:

gradient polyacrylamide gel electrophoresis

MG115:

Z–Leu–Leu–Nva–H (aldehyde)

REs:

root endopeptidase isoenzymes

TMB:

3 3′, 5, 5′–tetramethylbenzidine

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Acknowledgements

We thank Dr Kunpu Zhang for her kind gift of seeds used in this work. This work is also supported by the National Natural Science Foundation of China (Grant No. 30370851, 30871461).

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  1. College of Life Sciences, Nanjing Agricultural University, 210095, Nanjing, People’s Republic of China

    Ying Zhang, Junhua Zhong & Langlai Xu

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  1. Ying Zhang
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  2. Junhua Zhong
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  3. Langlai Xu
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Correspondence to Langlai Xu.

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Zhang, Y., Zhong, J. & Xu, L. Identification and biochemical characterization of 20S proteasome in wheat roots under salt stress. J. Plant Biochem. Biotechnol. 22, 62–70 (2013). https://doi.org/10.1007/s13562-012-0111-8

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  • DOI: https://doi.org/10.1007/s13562-012-0111-8

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