Cloning, expression and characterization of metalloproteinase HypZn from Aspergillus niger

Song, Peng; Xu, Wei; Wang, Kuiming; Zhang, Yang; Wang, Fei; Zhou, Xiuling; Hai-ying, Shi; Feng, Wei

doi:10.1371/journal.pone.0259809

Cited by 3 publications

(2 citation statements)

References 34 publications

(32 reference statements)

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“…For the carboxypeptidase assay, a peptide with an amino terminus blocked by benzyloxycarbonyl (CBZ; Fu et al, 2011 ; Song et al, 2021a ) or benzoyl (BZ; Ramirez Zavala et al, 2004 ; Heylen et al, 2010 ) is most commonly used as a substrate. Only carboxypeptidase can release amino acids from the carboxyl terminus.…”

Section: Detection Of Microbial Proteasesmentioning

confidence: 99%

Microbial proteases and their applications

Song,

Zhang,

Wang

et al. 2023

Front. Microbiol.

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Proteases (proteinases or peptidases) are a class of hydrolases that cleave peptide chains in proteins. Endopeptidases are a type of protease that hydrolyze the internal peptide bonds of proteins, forming shorter peptides; exopeptidases hydrolyze the terminal peptide bonds from the C-terminal or N-terminal, forming free amino acids. Microbial proteases are a popular instrument in many industrial applications. In this review, the classification, detection, identification, and sources of microbial proteases are systematically introduced, as well as their applications in food, detergents, waste treatment, and biotechnology processes in the industry fields. In addition, recent studies on techniques used to express heterologous microbial proteases are summarized to describe the process of studying proteases. Finally, future developmental trends for microbial proteases are discussed.

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Section: Detection Of Microbial Proteasesmentioning

confidence: 99%

Microbial proteases and their applications

Song,

Zhang,

Wang

et al. 2023

Front. Microbiol.

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“…Currently, cloning and expression of protease genes by genetic engineering technology are effective ways to identify novel proteases. To date, various proteases have been successfully expressed in Komagataella phaffii ( Pichia pastoris ) (Guo et al 2021 ; Mechri et al 2021 ; Song et al 2021 ). Microorganisms are the preferred sources of proteases, owing to their rapid growth, simple cultivation, and convenience for genetic manipulation (Mamo and Assefa 2018 ).…”

Section: Introductionmentioning

confidence: 99%

Characterization of a novel aspartic protease from Trichoderma asperellum for the preparation of duck blood peptides

Xue,

Yan,

et al. 2024

Appl Microbiol Biotechnol

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A novel aspartic protease gene (TaproA1) from Trichoderma asperellum was successfully expressed in Komagataella phaffii (Pichia pastoris). TaproA1 showed 52.8% amino acid sequence identity with the aspartic protease PEP3 from Coccidioides posadasii C735. TaproA1 was efficiently produced in a 5 L fermenter with a protease activity of 4092 U/mL. It exhibited optimal reaction conditions at pH 3.0 and 50 °C and was stable within pH 3.0–6.0 and at temperatures up to 45 °C. The protease exhibited broad substrate specificity with high hydrolysis activity towards myoglobin and hemoglobin. Furthermore, duck blood proteins (hemoglobin and plasma protein) were hydrolyzed by TaproA1 to prepare bioactive peptides with high ACE inhibitory activity. The IC50 values of hemoglobin and plasma protein hydrolysates from duck blood proteins were 0.105 mg/mL and 0.091 mg/mL, respectively. Thus, the high yield and excellent biochemical characterization of TaproA1 presented here make it a potential candidate for the preparation of duck blood peptides. Key points • An aspartic protease (TaproA1) from Trichoderma asperellum was expressed in Komagataella phaffii. • TaproA1 exhibited broad substrate specificity and the highest activity towards myoglobin and hemoglobin. • TaproA1 has great potential for the preparation of bioactive peptides from duck blood proteins.

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Cloning, Expression, and Characterization of a Metalloprotease from Thermophilic Bacterium Streptomyces thermovulgaris

Mushtaq,

Ahmed,

Mehmood

et al. 2024

Biology

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Proteases hydrolyze proteins and reduce them to smaller peptides or amino acids. Besides many biological processes, proteases play a crucial in different industrial applications. A 792 bp protease gene (nprB) from the thermophilic bacterium Streptomyces thermovulgaris was cloned and expressed in E. coli BL21 using pET 50b (+). Optimal recombinant protease expression was observed at 1 mM IPTG, 37 °C for 4 h. The resulting protease was observed in soluble form. The molecular mass estimated by SDS-PAGE and Western blot analysis of the protease (NprB) fused with His and Nus tag is ~70 KDa. The protease protein was purified by Ammonium sulfate precipitation and immobilized metal ion affinity chromatography. The optimum pH and temperature for protease activity using casein as substrate were 7.2 and 70 °C, respectively. The mature protease was active and retained 80% of its activity in a broad spectrum of pH 6–8 after 4 h of incubation. Also, the half-life of the protease at 70 °C was 4 h. EDTA (5 mM) completely inhibited the enzyme, proving the isolated protease was a metalloprotease. NprB activity was enhanced in the presence of Zn2+, Mn2+, Fe2+ and Ca2+, while Hg2+ and Ni2+ decreased its activity. Exposure to organic solvents did not affect the protease activity. The recombinant protease was stable in the presence of 10% organic solvents and surfactants. Further characterization showed that zinc-metalloprotease is promising for the detergent, laundry, leather, and pharmaceutical industries.

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Cloning, expression and characterization of metalloproteinase HypZn from Aspergillus niger

Cited by 3 publications

References 34 publications

Microbial proteases and their applications

Microbial proteases and their applications

Characterization of a novel aspartic protease from Trichoderma asperellum for the preparation of duck blood peptides

Cloning, Expression, and Characterization of a Metalloprotease from Thermophilic Bacterium Streptomyces thermovulgaris

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