Non-classical secretion of a type I L-asparaginase in Bacillus subtilis

Niu, Jiafeng; Meng, Fanqiang; Zhou, Yawen; Zhang, Chong; Lü, Fang; Chen, Meirong

doi:10.1016/j.ijbiomac.2021.03.104

Cited by 19 publications

(17 citation statements)

References 33 publications

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“…Because SignalP 3.0 predicted the most likely cleavage site (1–15 amino acids) of AGH17470 and the first and last 50 amino acids may contribute to secretion of nonclassical secretory proteins (Niu et al, 2021 ), seven truncated segments (amino acids 1–14, 1–50, 1–166, 167–215, 51–215, 15–215, and 1–215) were subjected to an alkaline phosphatase (PhoA) fusion assay for determining the key sequence involved in secretion. The results demonstrated that the first 50 and the last 50 amino acid sequences of AGH17470 are crucial for its secretion (Figure 1 ).…”

Section: Resultsmentioning

confidence: 99%

A prophage‐encoded nonclassical secretory protein of “Candidatus Liberibacter asiaticus” induces a strong immune response in Nicotiana benthamiana and citrus

Wang

Zeng

et al. 2022

Molecular Plant Pathology

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Huanglongbing (HLB), associated with “Candidatus Liberibacter asiaticus” (CLas), is a globally devastating plant disease. The highly reduced genome of CLas encodes a number of secretory proteins. The conserved prophage‐encoded protein AGH17470 is herein identified as a nonclassical secretory protein. We confirmed that the N‐terminal and C‐terminal sequences jointly determine the secretion of AGH17470. The transient expression of AGH17470 protein in Nicotiana benthamiana caused hypersensitive response (HR) cell death in infiltrated leaves and systemically infected leaves as well as the dwarfing of the entire plant, suggesting that AGH17470 is involved in the plant immune response, growth, and development. Overexpression of AGH17470 in N. benthamiana and citrus plants up‐regulated the transcription of pathogenesis‐related and salicylic acid (SA)‐signalling pathway genes and promoted SA accumulation. Furthermore, transient expression of AGH17470 enhanced the resistance of sweet orange to Xanthomonas citri subsp. citri. To our knowledge, AGH17470 is the first prophage‐encoded secretory protein demonstrated to elicit an HR and induce a strong plant immune response. The findings have increased our understanding of prophage‐encoded secretory protein genes, and the results provide clues as to the plant defence response against CLas.

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Section: Resultsmentioning

confidence: 99%

A prophage‐encoded nonclassical secretory protein of “Candidatus Liberibacter asiaticus” induces a strong immune response in Nicotiana benthamiana and citrus

Wang

Zeng

et al. 2022

Molecular Plant Pathology

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“…The amount of ammonia liberated from the hydrolysis of L-asparagine was calculated to determine L-asparaginase activity, and the BlAase activity was determined using the method described in our earlier study [ 22 ]. The amount of enzyme necessary to liberate 1 μmol of ammonia per mL and per min at pH 8.0 at 37 °C was established as one unit (U) of L-asparaginase activity.…”

Section: Methodsmentioning

confidence: 99%

“…Pang et al, for example, used a mix of promoter and RBS engineering methods to greatly improve pullulanase yield [ 21 ]. Previously, a type I L-asparaginase from Bacillus licheniformis Z-1 (BlAase) with good enzymatic characteristics was successfully produced and secreted in B. subtilis [ 22 ]. However, the yield of this enzyme was quite low, and thus hard to meet industrial demand.…”

Section: Introductionmentioning

confidence: 99%

Cis-Element Engineering Promotes the Expression of Bacillus subtilis Type I L-Asparaginase and Its Application in Food

Niu

Yan

Shen

et al. 2022

IJMS

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Type I L-asparaginase from Bacillus licheniformis Z-1 (BlAase) was efficiently produced and secreted in Bacillus subtilis RIK 1285, but its low yield made it unsuitable for industrial use. Thus, a combined method was used in this study to boost BlAase synthesis in B. subtilis. First, fifteen single strong promoters were chosen to replace the original promoter P43, with PyvyD achieving the greatest BlAase activity (436.28 U/mL). Second, dual-promoter systems were built using four promoters (PyvyD, P43, PaprE, and PspoVG) with relatively high BlAase expression levels to boost BlAase output, with the engine of promoter PaprE-PyvyD reaching 502.11 U/mL. The activity of BlAase was also increased (568.59 U/mL) by modifying key portions of the PaprE-PyvyD promoter. Third, when the ribosome binding site (RBS) sequence of promoter PyvyD was replaced, BlAase activity reached 790.1 U/mL, which was 2.27 times greater than the original promoter P43 strain. After 36 h of cultivation, the BlAase expression level in a 10 L fermenter reached 2163.09 U/mL, which was 6.2 times greater than the initial strain using promoter P43. Moreover, the application potential of BlAase on acrylamide migration in potato chips was evaluated. Results showed that 89.50% of acrylamide in fried potato chips could be removed when combined with blanching and BlAase treatment. These findings revealed that combining transcription and translation techniques are effective strategies to boost recombinant protein output, and BlAase can be a great candidate for controlling acrylamide in food processing.

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“…Type I L-ASNases are constitutively expressed enzymes localized in the cytoplasm and have a relatively high Km for L-asparagine. L-ASNases Bacillus subtilis [ 12 ], Pyrococcus horikoshii [ 13 ] and Acinetobacter soli [ 14 ] are the most studied examples of type I enzymes, which show a relatively low affinity to L-asparagine, resulting in nontherapeutic applications. Type II bacterial L-ASNases are periplasmic enzymes with induced expression during anaerobiosis that have a high affinity for L-asparagine and a wide substrate specificity, resulting in potent antitumor activity [ 15 ].…”

Section: Introductionmentioning

confidence: 99%

Molecular Analysis of L-Asparaginases for Clarification of the Mechanism of Action and Optimization of Pharmacological Functions

et al. 2022

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L-asparaginases (EC 3.5.1.1) are a family of enzymes that catalyze the hydrolysis of L-asparagine to L-aspartic acid and ammonia. These proteins with different biochemical, physicochemical and pharmacological properties are found in many organisms, including bacteria, fungi, algae, plants and mammals. To date, asparaginases from E. coli and Dickeya dadantii (formerly known as Erwinia chrysanthemi) are widely used in hematology for the treatment of lymphoblastic leukemias. However, their medical use is limited by side effects associated with the ability of these enzymes to hydrolyze L-glutamine, as well as the development of immune reactions. To solve these issues, gene-editing methods to introduce amino-acid substitutions of the enzyme are implemented. In this review, we focused on molecular analysis of the mechanism of enzyme action and to optimize the antitumor activity.

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Non-classical secretion of a type I L-asparaginase in Bacillus subtilis

Cited by 19 publications

References 33 publications

A prophage‐encoded nonclassical secretory protein of “Candidatus Liberibacter asiaticus” induces a strong immune response in Nicotiana benthamiana and citrus

A prophage‐encoded nonclassical secretory protein of “Candidatus Liberibacter asiaticus” induces a strong immune response in Nicotiana benthamiana and citrus

Cis-Element Engineering Promotes the Expression of Bacillus subtilis Type I L-Asparaginase and Its Application in Food

Molecular Analysis of L-Asparaginases for Clarification of the Mechanism of Action and Optimization of Pharmacological Functions

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