Abstract
Twelve peptides, including eight conservative amino acid residues in the amino acid sequence of hydrophilic S helix of the alternative oxidase (AOX), were synthesized by solid-phase method. The polypeptide was coupled with α-chymotrypsinogen, and the antibodies were obtained through immunizing domestic rabbit by injecting this complex. By using these antibodies, which were raised to immunoreact with total proteins of purified mitochondria from different organs of mung bean seedlings, we find that there are two hybridizable AOX bands in mitochondria. Their molecular weights are about 35 and 38 ku, respectively. Moreover, the respiratory parameters of hypocotyl, true leaf and cotyledon of mung bean seedlings show that true leaf has the highest total respiration (Vt), alternative pathway (AP) capacity (Valt) and the activity of AP (ρValt) among the three organs. Vt andρV alt of cotyledon ranked the second. Hypocotyl has the lowest Vt andρV alt, but its Valt is higher than that of cotyledon. These results are consistent with the analysis of Western blotting of expression of AOX. The highest Vt andρV alt in true leaf are accompanied two hybridizable polypeptides of AOX protein, 35 ku and 38 ku respectively. The next is cotyledon Vt andρV alt with only one 38 ku hybridizable polypeptide of AOX protein. HypocotylV t andρV alt is the lowest and its immunoblotting band is similar to that of cotyledon, but the expressive amount of 38 ku protein is less than that of cotyledon. The results suggest that the 35 ku AOX may contribute mainly to true leafρV alt.
Similar content being viewed by others
Abbreviations
- AOX:
-
alternative oxidase
- AP:
-
alternative pathway
- FW:
-
fresh weight
- SHAM:
-
salicylhydroxamic acid
References
Liang, Z., Liang, H. G., The respiratory metabolism of plants, in Plant Physiology and Molecular Biology (eds, Yu, S. W., Tang, Z. C.) (in Chinese), 2nd ed., Beijing: Science Press, 1998, 344–365.
Lü, C. S., Liang, H. G., Induced respiration in melon fruits, Scientia Sinica, 1963,12(4): 616.
Liang, H. G., Lü, C. S., A comparative study of CN-resistant respiration in different cultures of tobacco callus, Plant Physiol., 1984,75: 876.
Elthon, T. E., McIntosh, L., Identification of the alternative terminal oxidase of higher plant mitochondria, Proc. Natl. Acad. Sci. USA, 1987,84: 8399.
Elthon, T. E., Nickels, R. L., McIntosh, L., Monoclonal antibodies to the alternative oxidase of higher plant mitochondria, Plant Physiol., 1989,89: 1311.
Liang, W. S., Liang, H. G., Progress of the alternative oxidase, Chinese Bulletin of Botany (in Chinese), 1997,14{(sn2)}: 9.
Liang, W. S., Liang, H. G., Induction of alternative oxidase expression by endogenous ethylene in aging potato slices, Acta Phytophysiol. Sin. (in Chinese), 1999,25{(sn2)}: 205.
He, J. X., Wei, Z. Q., Liang, H. G., Effects of water stress on development, and operation and gene expression of cyanide-resistant respiratory pathway in wheat, Science in China, Ser. C, 1999,42{(sn3)}: 300.
McIntosh, L., Molecular biology of the alternative oxidase, Plant Physiol., 1994,105: 781.
Wang, J., Zhang, L. X., Liu, Z. L. et al., A possible calcium binding site in D1 protein: A fluorescence and FTIR study of the interaction between lanthanides and a synthetic peptide, Photosynthesis Research, 1995,44: 297.
Li, X. P., Du, L. F., Liang, H. G. et al., Preparation and identification of antidodecapeptide ofpolypeptide D1 orphotosystem II reaction center, Prog. Biochem. Biophys. (in Chinese), 1997,24{(sn3)}: 283.
Wen, J. Q., Liang, H. G., Studies on energy status and mitochondria respiration during growth and senescence of mung bean cotyledons, Physiol. Plant, 1993,89: 805.
Laemmli, U. K., Cleavage of structural proteins during the assembly of the head of bacteriophage T4, Nature, 1970,227: 680.
Sambrook, J., Fritsch, E. F., Mariatis, T., Molecular Cloning: A Laboratory Manual (translated by Jin, D. Y., Li, M. F.) (in Chinese), 2nd ed., Beijing: Science Press, 1989, 888–898.
Bingham, I. J., Farrar, J. F., Activity and capacity of respiratory pathway in barley roots deprived of inorganic nutrients, Plant Physiol. Biochem., 1989,27: 847.
Rhoads, D. M., McIntosh, L., Isolation and characterization of a cDNA clone encoding an alternative oxidase protein ofSauromatum guttatum, Proc. Natl. Acad. Sci. USA, 1991,88: 2122.
Vanlerberghe, G. C., McIntosh, L., Alternative oxidase: From gene to function, Annu. Rev. Plant Physiol. Plant Mol. Biol., 1997,48: 703.
Young, C. R., Atassi, M. Z., Antibodies to sperm-whale myoglobin evoked by free synthetic peptides of an antigenic site, Immunol. Commun., 1982,11{(sn1)}: 9.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Li, C., Liang, H., Du, L. et al. Studies on the relationship between cyanide-resistant respiration and expression of alternative oxidase in mung bean using antibodies prepared by synthetic polypeptide. Sci. China Ser. C.-Life Sci. 44, 66–72 (2001). https://doi.org/10.1007/BF02882074
Received:
Issue Date:
DOI: https://doi.org/10.1007/BF02882074