Comparative Analysis of Genomics and Proteomics in Bacillus thuringiensis 4.0718

Abstract

Bacillus thuringiensis is a widely used biopesticide that produced various insecticidal active substances during its life cycle. Separation and purification of numerous insecticide active substances have been difficult because of the relatively short half-life of such substances. On the other hand, substances can be synthetized at different times during development, so samples at different stages have to be studied, further complicating the analysis. A dual genomic and proteomic approach would enhance our ability to identify such substances, and particularily using mass spectrometry-based proteomic methods. The comparative analysis for genomic and proteomic data have showed that not all of the products deduced from the annotated genome could be identified among the proteomic data. For instance, genome annotation results showed that 39 coding sequences in the whole genome were related to insect pathogenicity, including five cry genes. However, Cry2Ab, Cry1Ia, Cytotoxin K, Bacteriocin, Exoenzyme C3 and Alveolysin could not be detected in the proteomic data obtained. The sporulation-related proteins were also compared analysis, results showed that the great majority sporulation-related proteins can be detected by mass spectrometry. This analysis revealed Spo0A~P, SigF, SigE(+), SigK(+) and SigG(+), all known to play an important role in the process of spore formation regulatory network, also were displayed in the proteomic data. Through the comparison of the two data sets, it was possible to infer that some genes were silenced or were expressed at very low levels. For instance, found that cry2Ab seems to lack a functional promoter while cry1Ia may not be expressed due to the presence of transposons. With this comparative study a relatively complete database can be constructed and used to transform hereditary material, thereby prompting the high expression of toxic proteins. A theoretical basis is provided for constructing highly virulent engineered bacteria and for promoting the application of proteogenomics in the life sciences.

A Genomic and Proteomic Approach to Identify and Quantify the Expressed Bacillus thuringiensis Proteins in the Supernatant and Parasporal Crystal

“…The quantity of the proteins expressed at the three different growth phases for the Bt isolates E-SE10.2 and O-V84.2 were 10.2–3.5% and 10.4–8.2%, 8.3–2.8%, respectively, of their genome encoded sequences. This low percentage of expressed proteins detected indicates that, in our experimental conditions, we only detect a small part of the predicted proteins by the genome data prediction, a phenomenon that has also been found in other studies [ 21 , 22 ]. The low percentage of detected expressed proteins should not be interpreted as that the rest of the proteins cannot be expressed, since they could do it under different growth conditions.…”

“…On the other hand, the development of the mass spectrometry (MS)-based proteomics has enabled the detection of proteins from complex mixtures from different stages of a microorganism [ 18 ]. The combination of the genomic and proteomic approaches is a very successful approach for validation and correction of predicted genomic coding information in a wide variety of organisms [ 18 , 19 , 20 , 21 , 22 ].…”

Proteomic analysis of the influence of Cu2+ on the crystal protein production of Bacillus thuringiensis X022

“…B. thuringiensis X022 cultivated in original fermentation medium and 10 −6 mol/L Cu 2+ added fermentation medium were harvested from three replicates and mixed together at the time point of 44 h cultivation. The procedure of proteins extraction and trypsin digestion was carried out as previously described [ 34 , 35 ]. Briefly, after washed with PBS buffer (10 mM, pH 7.8), The cells were mechanically disrupted with disposable grinding pestles, and then treated with ultrasonication (SCIENTZ 98-III) for 10 min at 4 °C.…”

Comparative Analysis of Genomics and Proteomics in the New Isolated Bacillus thuringiensis X022 Revealed the Metabolic Regulation Mechanism of Carbon Flux Following Cu2+ Treatment

“…Since the first complete whole genome sequencing of Bt strain serovar konkukian 97-27 in 2004 ( Han et al, 2006 ), the number of sequenced genomes has increased sharply over the past decade, among which, 15 Bt strains have been completely sequenced up to now. The whole 6.3 Mb genome of Bt 4.0718 has been sequenced by our laboratory ( Rang et al, 2015 ). It comprises a circular chromosome of 5.6 Mb encoding 6747 open reading frames (ORFs) and seven circular plasmids.…”