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Detection of Toxin Proteins from Bacillus thuringiensis Strain 4.0718 by Strategy of 2D-LC–MS/MS

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Abstract

Bacillus thuringiensis is a kind of insecticidal microorganism which can produce a variety of toxin proteins, it is particularly important to find an effective strategy to identify novel toxin proteins rapidly and comprehensively with the discovery of the wild-type strains. Multi-dimensional high-performance liquid chromatography combined with mass spectrometry has become one of the main methods to detect and identify toxin proteins and proteome of B. thuringiensis. In this study, protein samples from B. thuringiensis strain 4.0718 were analyzed on the basis of two-dimensional liquid chromatography–tandem mass spectrometry (2D-LC–MS/MS), and tryptic peptides of whole cell from the late sporulation phase were eluted at different concentration gradients of ammonium chloride and followed by secondary mass spectrum identification. 831 and 894 proteins were identified from two biological replicates, respectively, while 1,770 and 1,859 peptides were detected correspondingly. Among the identified proteins and peptides, 606 proteins and 1,259 peptides were detected in both replicates, which mean that 1,119 proteins and 2,370 peptides were unique to the proteome of this strain. A total of 15 toxins have been identified successfully, and seven of them were firstly discovered in B. thuringiensis strain 4.0718 that were Crystal protein (A1E259), pesticidal protein (U5KS09), Cry2Af1 (A4GVF0), Cry2Ad (Q9RM89), Cry1 (K4HMB5), Cry1Bc (Q45774), and Cry1Ga (Q45746). The proteomic strategy employed in the present study has provided quick and exhaustive identification of toxins produced by B. thuringiensis.

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Acknowledgments

This work was supported by the National Natural Science Foundation of China (31370116), the National Basic Research Program (973) of China (2012CB722301), the National High Technology Research and Development program (863) of China (2011AA10A203), the Key Project of Hunan Provincial Education Department (13CY002, 10CY013), and the Cooperative Innovation Center of Engineering and New Products for Developmental Biology of Hunan Province (20134486).

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Authors and Affiliations

  1. Hunan Provincial Key Laboratory of Microbial Molecular Biology-State Key laboratory Breeding Base of Microbial Molecular Biology, College of Life Science, Hunan Normal University, Changsha, 410081, China

    Qi Yang, Sijia Tang, Jie Rang, Mingxing Zuo, Xuezhi Ding, Yunjun Sun, Pinghui Feng & Liqiu Xia

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  1. Qi Yang
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  2. Sijia Tang
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  3. Jie Rang
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  8. Liqiu Xia
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Correspondence to Liqiu Xia.

Electronic Supplementary Material

Below is the link to the electronic supplementary material.

284_2014_747_MOESM1_ESM.xls

Identification of Bacillus thuringiensis proteins from each biological replicates. Supplementary material 1 (XLS 282 kb)

284_2014_747_MOESM2_ESM.xls

Identification of Bacillus thuringiensis peptides from each biological replicates. Supplementary material 2 (XLS 352 kb)

284_2014_747_MOESM3_ESM.doc

Unique peptide and their consecutive b- or y-ions figures of identified toxin proteins during detection. Supplementary material 3 (DOC 1738 kb)

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Yang, Q., Tang, S., Rang, J. et al. Detection of Toxin Proteins from Bacillus thuringiensis Strain 4.0718 by Strategy of 2D-LC–MS/MS. Curr Microbiol 70, 457–463 (2015). https://doi.org/10.1007/s00284-014-0747-9

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  • DOI: https://doi.org/10.1007/s00284-014-0747-9

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