Dataset Information

Novel Aflatoxin-Degrading Enzyme from Bacillus shackletonii L7.

ABSTRACT: Food and feed contamination by aflatoxin (AF)B? has adverse economic and health consequences. AFB? degradation by microorganisms or microbial enzymes provides a promising preventive measure. To this end, the present study tested 43 bacterial isolates collected from maize, rice, and soil samples for AFB?-reducing activity. The higher activity was detected in isolate L7, which was identified as Bacillus shackletonii. L7 reduced AFB?, AFB?, and AFM? levels by 92.1%, 84.1%, and 90.4%, respectively, after 72 h at 37 °C. The L7 culture supernatant degraded more AFB? than viable cells and cell extracts; and the degradation activity was reduced from 77.9% to 15.3% in the presence of proteinase K and sodium dodecyl sulphate. A thermostable enzyme purified from the boiled supernatant was designated as Bacillus aflatoxin-degrading enzyme (BADE). An overall 9.55-fold purification of BADE with a recovery of 39.92% and an activity of 3.85 × 10³ U·mg-1 was obtained using chromatography on DEAE-Sepharose. BADE had an estimated molecular mass of 22 kDa and exhibited the highest activity at 70 °C and pH 8.0, which was enhanced by Cu2+ and inhibited by Zn2+, Mn2+, Mg2+, and Li?. BADE is the major protein involved in AFB? detoxification. This is the first report of a BADE isolated from B. shackletonii, which has potential applications in the detoxification of aflatoxins during food and feed processing.

SUBMITTER: Xu L

PROVIDER: S-EPMC5308268 | biostudies-literature | 2017 Jan

REPOSITORIES: biostudies-literature

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Novel Aflatoxin-Degrading Enzyme from Bacillus shackletonii L7.

Xu Liang L Eisa Ahmed Mohamed Farah MF Sangare Lancine L Zhao Yueju Y Selvaraj Jonathan Nimal JN Xing Fuguo F Wang Yan Y Yang Hongping H Liu Yang Y

Toxins 20170114 1

Food and feed contamination by aflatoxin (AF)B₁ has adverse economic and health consequences. AFB₁ degradation by microorganisms or microbial enzymes provides a promising preventive measure. To this end, the present study tested 43 bacterial isolates collected from maize, rice, and soil samples for AFB₁-reducing activity. The higher activity was detected in isolate L7, which was identified as <i>Bacillus shackletonii</i>. L7 reduced AFB₁, AFB₂, and AFM₁ levels by 92.1%, 84.1%, and 90.4%, respect ...[more]

PMID: 28098812

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Project description:Although ADAMTS-4 and ADAMTS-5 are the principal aggrecanases in mice and humans, mice lacking both these enzymes (ADAMTS-4/-5Dcat) are healthy, viable, with no overt skeletal phenotype, suggesting the presence of an alternative aggrecanase in these mice. We previously identified a novel aggrecanase activity that was regulated by retinoic acid, but not IL-1a. The upregulated activity cleaved aggrecan at E↓A bonds. This study aimed to identify the alternative aggrecanase. Femoral head cartilage from TS-4/5Dcat mice was stimulated with IL-1a or retinoic acid and total RNA was analysed by microarray. Candidate, upregulated genes identified by Quantile Normalisation included ADAMTS, MMPm cathepsin and calpain families as putative aggrecanases. Three criteria, including quantitative RT-PCR analyses, sensitivity to retinoic acid treatment and sensitivity to TIMP-3 inhibition, were then used as readouts to identify candidate extracellular proteinases whose mRNA expression levels were increased by retinoic acid, but not by IL-1a treatment. Thereafter, gene silencing was used to identify the most likely aggrecanase in chondrocytes from TS-4/5Dcat mice. Aggrecanase activity was monitored by Western blotting and immunohistochemistry with neoepitope antibodies. The microarray analyses identified ADAMTS-9, MMP-11 and calpain-5 as candidate aggrecanases that were upregulated by retinoic acid. While qPCR confirmed this finding, calpain 5 was excluded from the candidate list because it was not inhibited by TIMP-3. MMP-11 was also excluded because it was upregulated by IL-1a. ADAMTS-9 therefore emerged as a candidate for the novel aggrecanase. Gene silencing confirmed ADAMTS-9 as the putative aggrecanase. Immunohistochemistry revealed that ADAMTS-9 in mouse knee joints was expressed in the proliferative zone of both wildtype and TS-4/5Dcat growth plates, and also in the calcified cartilage of the wildtype hypertrophic zone. In conclusion, ADAMTS-9 is an aggrecanase that cleaves at FREEE1467↓1468GLGS in the chondroitin sulphate-rich region of aggrecan.

Dataset Information

Novel Aflatoxin-Degrading Enzyme from Bacillus shackletonii L7.

Publications

Novel Aflatoxin-Degrading Enzyme from Bacillus shackletonii L7.

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