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Identification of B6T173 (ZmPrx35) as the prevailing peroxidase in highly insect-resistant maize (Zea mays, p84C3) kernels by activity-directed purification.

López-Castillo LM, López-Arciniega JA, Guerrero-Rangel A, Valdés-Rodríguez S, Brieba LG, García-Lara S, Winkler R - Front Plant Sci (2015)

Bottom Line: Contrary to their biological importance, only very few plant PODs have been proven on protein level, because their low abundance makes them difficult to detect in standard proteomics work-flows.A statistically significant positive correlation between POD activity and post-harvest insect resistance has been found for maize (Zea mays, p84C3) kernels.Our findings support the screening for insect resistant maize variants and the construction of genetically optimized maize plants.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Biochemical and Instrumental Analysis, Department of Biotechnology and Biochemistry, Cinvestav Unidad Irapuato Irapuato, Mexico ; Laboratorio Nacional de Genómica para la Biodiversidad, Unidad de Genómica Avanzada del Centro de Investigación y de Estudios Avanzados - Instituto Politécnico Nacional Irapuato, Mexico.

ABSTRACT
Plant peroxidases (PODs) are involved in diverse physiological processes, including defense against pathogens and insects. Contrary to their biological importance, only very few plant PODs have been proven on protein level, because their low abundance makes them difficult to detect in standard proteomics work-flows. A statistically significant positive correlation between POD activity and post-harvest insect resistance has been found for maize (Zea mays, p84C3) kernels. In combining activity-directed protein purification, genomic and proteomic tools we found that protein B6T173 (ZmPrx35) is responsible for the majority of the POD activity of the kernel. We successfully produced recombinant ZmPrx35 protein in Escherichia coli and demonstrate both, in vitro activity and the presence of a haem (heme) cofactor of the enzyme. Our findings support the screening for insect resistant maize variants and the construction of genetically optimized maize plants.

No MeSH data available.


Related in: MedlinePlus

B6T173 (ZmPrx35) iron cofactor detection. (A) Spectra of absorbance (200-800 nm) of ZmPrx35 purified fractions, and of a negative control (At CytoTPI). The arrow shows the peak of absorbance at 450 nm (Soret band). (B) Quantitative luminol assay of ZmPrx35 and AtCyto TPI (as a negative control) under native and unfolded conditions.
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Figure 5: B6T173 (ZmPrx35) iron cofactor detection. (A) Spectra of absorbance (200-800 nm) of ZmPrx35 purified fractions, and of a negative control (At CytoTPI). The arrow shows the peak of absorbance at 450 nm (Soret band). (B) Quantitative luminol assay of ZmPrx35 and AtCyto TPI (as a negative control) under native and unfolded conditions.

Mentions: UV-VIS spectra of recombinant ZmPrx35 showed increased absorbance with a maximum at about 450 nm (Figure 5A), which corresponds to a Soret-band feature. Thus, the spectroscopic data indicate the presence a haem group, which is a main structural characteristic of PODs (Dalton et al., 1996; Shannon et al., 1966) and related haem-dependent redox proteins such as catalase (Ray et al., 2012).


Identification of B6T173 (ZmPrx35) as the prevailing peroxidase in highly insect-resistant maize (Zea mays, p84C3) kernels by activity-directed purification.

López-Castillo LM, López-Arciniega JA, Guerrero-Rangel A, Valdés-Rodríguez S, Brieba LG, García-Lara S, Winkler R - Front Plant Sci (2015)

B6T173 (ZmPrx35) iron cofactor detection. (A) Spectra of absorbance (200-800 nm) of ZmPrx35 purified fractions, and of a negative control (At CytoTPI). The arrow shows the peak of absorbance at 450 nm (Soret band). (B) Quantitative luminol assay of ZmPrx35 and AtCyto TPI (as a negative control) under native and unfolded conditions.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4553411&req=5

Figure 5: B6T173 (ZmPrx35) iron cofactor detection. (A) Spectra of absorbance (200-800 nm) of ZmPrx35 purified fractions, and of a negative control (At CytoTPI). The arrow shows the peak of absorbance at 450 nm (Soret band). (B) Quantitative luminol assay of ZmPrx35 and AtCyto TPI (as a negative control) under native and unfolded conditions.
Mentions: UV-VIS spectra of recombinant ZmPrx35 showed increased absorbance with a maximum at about 450 nm (Figure 5A), which corresponds to a Soret-band feature. Thus, the spectroscopic data indicate the presence a haem group, which is a main structural characteristic of PODs (Dalton et al., 1996; Shannon et al., 1966) and related haem-dependent redox proteins such as catalase (Ray et al., 2012).

Bottom Line: Contrary to their biological importance, only very few plant PODs have been proven on protein level, because their low abundance makes them difficult to detect in standard proteomics work-flows.A statistically significant positive correlation between POD activity and post-harvest insect resistance has been found for maize (Zea mays, p84C3) kernels.Our findings support the screening for insect resistant maize variants and the construction of genetically optimized maize plants.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Biochemical and Instrumental Analysis, Department of Biotechnology and Biochemistry, Cinvestav Unidad Irapuato Irapuato, Mexico ; Laboratorio Nacional de Genómica para la Biodiversidad, Unidad de Genómica Avanzada del Centro de Investigación y de Estudios Avanzados - Instituto Politécnico Nacional Irapuato, Mexico.

ABSTRACT
Plant peroxidases (PODs) are involved in diverse physiological processes, including defense against pathogens and insects. Contrary to their biological importance, only very few plant PODs have been proven on protein level, because their low abundance makes them difficult to detect in standard proteomics work-flows. A statistically significant positive correlation between POD activity and post-harvest insect resistance has been found for maize (Zea mays, p84C3) kernels. In combining activity-directed protein purification, genomic and proteomic tools we found that protein B6T173 (ZmPrx35) is responsible for the majority of the POD activity of the kernel. We successfully produced recombinant ZmPrx35 protein in Escherichia coli and demonstrate both, in vitro activity and the presence of a haem (heme) cofactor of the enzyme. Our findings support the screening for insect resistant maize variants and the construction of genetically optimized maize plants.

No MeSH data available.


Related in: MedlinePlus