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Identity of an ABA-activated 46 kDa mitogen-activated protein kinase from Zea mays leaves: partial purification, identification and characterization

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Abstract

Mitogen-activated protein kinase (MAPK) cascades have been shown to be important components in abscisic acid (ABA) signal transduction pathway. In this study, a 46 kDa MAPK (p46MAPK) induced by ABA was partially purified from maize (Zea mays) by Q-Sepharose FF, Phenyl-Sepharose FF, Resource Q, Mono QTM 5/50 GL, poly-l-lysine-agarose, and Superdex 75 prep-grade columns, and was identified as ZmMAPK5 (gi|4239889) by the matrix-assisted laser desorption/ionization time-of-flight/time-of-flight (MALDI-TOF/TOF) mass spectrometry. Furthermore, the kinase showed optimal activity at pH 8.0, 30°C, and 10 mM MgCl2; the K m for myelin basic protein (MBP) substrate and ATP were 0.13 μg μl−1 and 62 μM, respectively. MBP was the preferred substrate, of which the threonine residue was phosphorylated. Finally, the kinase was found to respond to diverse extracelluar stimuli. These results enable us to further reveal the function of the ZmMAPK5 in ABA signaling.

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Abbreviations

ABA:

Abscisic acid

MALDI-TOF/TOF:

Matrix-assisted laser desorption/ionization time-of-flight/time-of-flight

MAPK:

Mitogen-activated protein kinase

MAPKK:

Mitogen-activated protein kinase kinase

MAPKKK:

Mitogen-activated protein kinase kinase kinase

MBP:

Myelin basic protein

MS:

Mass spectrum

SA:

Salicylic acid

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Acknowledgments

This work was supported by the National Natural Science Foundation of China (grant nos. 30571122 and 30671247 to M. J.), the Universities Qing-Lan Project of Jiangsu Province (to M. J.), the Graduate Research and Innovation Plan of Jiangsu Province (grant no. CX07B_052Z to H. D.), and the Open Project of the National Key Laboratory of Crop Genetics and Germplasm Enhancement of Nanjing Agricultural University (grant no. ZW2007002 to M. J.).

Author information

Correspondence to Mingyi Jiang.

Additional information

The authors Haidong Ding and Aying Zhang contributed equally to the work.

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Ding, H., Zhang, A., Wang, J. et al. Identity of an ABA-activated 46 kDa mitogen-activated protein kinase from Zea mays leaves: partial purification, identification and characterization. Planta 230, 239–251 (2009). https://doi.org/10.1007/s00425-009-0938-y

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Keywords

  • Abscisic acid
  • Mass spectrometry
  • Mitogen-activated protein kinase
  • Protein purification
  • Zea