Improved catalytic efficiency, thermophilicity, anti-salt and detergent tolerance of keratinase KerSMD by partially truncation of PPC domain

Fang, Zhen; Zhang, Juan; Du, Guocheng; Chen, Jian

doi:10.1038/srep27953

Cited by 31 publications

(22 citation statements)

References 39 publications

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“…Recent studies have focused on protein engineering to improve the catalytic efficiency, thermophilicity, anti-salt, and detergent tolerance of keratinase for industry use 30, 35, 36 . Our phylogenetic and structural analysis (Fig.…”

Section: Discussionmentioning

confidence: 99%

The discovery of novel heat-stable keratinases from Meiothermus taiwanensis WR-220 and other extremophiles

Chen

et al. 2017

Sci Rep

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Billions of tons of keratin bio-wastes are generated by poultry industry annually but discarded that result in serious environmental pollution. Keratinase is a broad spectrum protease with the unique ability to degrade keratin, providing an eco-friendly way to convert keratin wastes to valuable amino acids. In this report, a feather-degrading thermophilic bacterium, Meiothermus taiwanensis WR-220, was investigated due to its ability to apparently complete feather decay at 65 °C in two days. By genomics, proteomics, and biochemical approaches, the extracellular heat-stable keratinase (MtaKer) from M. taiwanensis WR-220 was identified. The recombinant MtaKer (rMtaKer) possesses keratinolytic activities at temperatures ranging from 25 to 75 °C and pH from 4 to 11, with a maximum keratinolytic activity at 65 °C and pH 10. The phylogenetic and structural analysis revealed that MtaKer shares low sequence identity but high structural similarity with known keratinases. Accordingly, our findings have enabled the discovery of more keratinases from other extremophiles, Thermus and Deinococcus. Proteins encoded in the extremophiles shall be evolved to be functional in the extreme conditions. Hence, our study expands the current boundary of hunting keratinases that can tolerate extreme conditions for keratin wastes biorecycle and other industrial applications.

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Section: Discussionmentioning

confidence: 99%

The discovery of novel heat-stable keratinases from Meiothermus taiwanensis WR-220 and other extremophiles

Chen

et al. 2017

Sci Rep

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“…Usually, keratinase activities are inhibited, stabilized, or enhanced in a solution that contains surfactants, reducing agents, or solvents. These variable behaviors of keratinases toward chemical agents are attributable to the nature of their side chains/interaction patterns which allosterically regulate the structural orientation and biocatalytic efficiency of the enzymes (Fang et al, 2016). The activity of the alkaline keratinase from S. aureofaciens K13 was enhanced when incubated with SDS, Tween-80 or Triton-X100 (Gong et al, 2015).…”

Section: Biochemical Properties Of Keratinasementioning

confidence: 99%

Microbial Keratinase: Next Generation Green Catalyst and Prospective Applications

et al. 2020

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The search for novel renewable products over synthetics hallmarked this decade and those of the recent past. Most economies that are prospecting on biodiversity for improved bio-economy favor renewable resources over synthetics for the potential opportunity they hold. However, this field is still nascent as the bulk of the available resources are non-renewable based. Microbial metabolites, emphasis on secondary metabolites, are viable alternatives; nonetheless, vast microbial resources remain under-exploited; thus, the need for a continuum in the search for new products or bio-modifying existing products for novel functions through an efficient approach. Environmental distress syndrome has been identified as a factor that influences the emergence of genetic diversity in prokaryotes. Still, the process of how the change comes about is poorly understood. The emergence of new traits may present a high prospect for the industrially viable organism. Microbial enzymes have prominence in the bio-economic space, and proteases account for about sixty percent of all enzyme market. Microbial keratinases are versatile proteases which are continuously gaining momentum in biotechnology owing to their effective bio-conversion of recalcitrant keratin-rich wastes and sustainable implementation of cleaner production. Keratinase-assisted biodegradation of keratinous materials has revitalized the prospects for the utilization of cost-effective agro-industrial wastes, as readily available substrates, for the production of high-value products including amino acids and bioactive peptides. This review presented an overview of keratin structural complexity, the potential mechanism of keratin biodegradation, and the environmental impact of keratinous wastes. Equally, it discussed microbial keratinase; vis-à-vis sources, production, and functional properties with considerable emphasis on the ecological implication of microbial producers and catalytic tendency improvement strategies. Keratinase applications and prospective high-end use, including animal hide processing, detergent formulation, cosmetics, livestock feed, and organic fertilizer production, were also articulated.

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“…Since keratin is an insoluble protein, the study and application of keratinases are of particular interest. Some researchers have been devoted to purifying and characterizing the newly-found enzymes from various microorganisms [ 14 , 17 – 20 ], some searching for the suitable expression system for heterologous expression of keratinses [ 4 , 6 , 7 , 10 ], and some others made efforts to improve the activity of the protease [ 21 , 22 ]. For heterologous expression, E .…”

Section: Discussionmentioning

confidence: 99%

Secretory expression and purification of Bacillus licheniformis keratinase in insect cells

Huang¹,

Chen

Ren³

2017

PLoS ONE

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The keratinase (kerA) gene from Bacillus licheniformis PWD-1 was expressed and purified in insect cells. First, the sequence encoding Ker-His-Flag was designed based on the amino acid sequence of the protein and peptide and codon optimization in order to ensure the high expression in insect cells. In the next step, the synthetic DNA was inserted into the pUC57 vector and then sub-cloned in the pFastBac™-1 donor vector by BamHI/HindIII restriction sites. The constructed vector was transformed to E. coli DH10Bac™ cell to generate recombinant bacmid carrying Ker-His-Flag. Recombinant viruses were produced by infecting insect Spodoptera frugiperda (Sf9) cells with bacmid DNA and used for proteins production. Target proteins were purified from the cell supernatants by Ni2+-NTA affinity chromatography and evaluated by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and western blot. The purified product contained two peptides with molecular weights of 38 kDa and 30 kDa and had an optimal pH and temperature at 8.0 and 45°C for keratinolytic activity, respectively. The final product had a specific activity of about 635 U/mg. In summary, we have demonstrated that the open reading frame containing recombinant Ker-His-Flag was expressed and secreted by leader peptide of mellittin from Apis mellitera in insect cells and affinity purification through 8His-Flag tag. It presents an alternative technology for producing keratinases. To our knowledge, it was the first report on the expression of functional keratinase from Bacillus licheniformis in insect cells system.

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Improved catalytic efficiency, thermophilicity, anti-salt and detergent tolerance of keratinase KerSMD by partially truncation of PPC domain

Cited by 31 publications

References 39 publications

The discovery of novel heat-stable keratinases from Meiothermus taiwanensis WR-220 and other extremophiles

The discovery of novel heat-stable keratinases from Meiothermus taiwanensis WR-220 and other extremophiles

Microbial Keratinase: Next Generation Green Catalyst and Prospective Applications

Secretory expression and purification of Bacillus licheniformis keratinase in insect cells

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