Ficin: A protease extract with relevance in biotechnology and biocatalysis

As a result of issues raised by way of animal or microbial recombinant proteases, using proteases from vegetable origin is changing into more and more in style. Amongst them, sulfidryl proteases have a particular curiosity. Fiction is an excellent instance of this type of protease.
This paper goals to be to make a complete evaluation of the current makes use of of this enzyme, together with for instance protein hydrolysis, the manufacturing of bioactive peptides and antibodies fragments (researchers level that ficin outcomes are extra reproducible than utilizing different proteases), meat tenderization, milk coagulations in cheese making or peptide synthesis.

Efforts to get industrial immobilized biocatalysts of the enzyme will likely be additionally described. The evaluation reveals the huge potential and sensible prospect that this enzyme can have within the close to future.

Key phrases: Bioactive peptides; Ficin; Protease immobilization; Proteases; Proteolysis; Synthesis of peptides.

An alternate for proteinase Ok-heat-sensitive protease from fungus Onygena corvina for biotechnology: cloning, engineering, expression, characterization and particular utility for protein sequencing


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Background: A impartial, heat-sensitive serine protease (NHSSP) originating from the feather-degrading fungus Onygena corvina (O. corvina) was described and outlined as an alkaline serine protease of the subtilisin kind S8 household, exhibiting an enzymatic exercise at impartial pH. Usually, broad specificity proteases, akin to proteinase Ok or trypsin, have discovered quite a few purposes in analysis and biotechnology.
Outcomes: We report the cloning and expression within the yeast PichiaPink™ system, in addition to purification, and characterization of the NHSSP. Recombinant, His6-tagged NHSSP was effectively expressed from an optimized, artificial gene and purified utilizing a easy protocol based mostly on ammonium sulfate fractionation and hydrophobic interplay chromatography.
  • The enzyme reveals atypical C-terminal processing, the coded preproprotein undergoes sign peptide removing and maturation by means of the clipping of a propeptide part and 10 amino acids (aa) from the C-terminus, together with the His6-tag.
  • The deletion variant has been constructed, devoid of the C-terminal ORF section, thus eliminating the necessity for C-terminal processing. Each NHSSP variants exhibit very related enzymatic traits.
  • The purified enzymes have been characterised to find out the optimum proteolytic circumstances. We revealed that the mature NHSSP is reproducibly lively over a large pH vary from impartial to delicate acidic (pH of to eight.5), with an optimum at pH 6.8, and at temperatures of 15 to 50 °C with an optimum at 38-42 °C.
  • Apparently, we demonstrated that the protease could be totally deactivated by a average enhance in temperature of about 15 °C from the optimum to over 50 °C.
  • The protease was partially delicate to serine protease inhibitors, and never inhibited by chelating or decreasing brokers and detergents. SDS induced autolysis of NHSSP, which factors to a excessive stimulation of its proteolytic exercise.

Conclusions: The NHSSP was produced as a recombinant protein with excessive effectivity. In comparison with proteinase Ok, the commonest serine protease used, NHSSP reveals an approx. twofold increased particular exercise.

Protein sequencing is usually a beneficial technical utility for the protease. The protein protection is considerably increased compared to trypsin and reaches about 84-100% for β-lactoglobulin (BLG), antibody (mAb) mild and heavy chains.

Moreover, the choice to carry out digestions at impartial to barely acidic pH-values right down to pH avoids modification of peptides, e.g. because of deamidation.

Key phrases: Expression; Meals processing; Onygena corvina; Pichia pastoris; Protease; Protein degradation; Protein sequencing; Proteinase Ok; Serine protease; Waste processing.
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Bioactive molecules from protists: Views in biotechnology

For a whole bunch of years, mankind has benefited from the pure metabolic processes of microorganisms to acquire primary merchandise akin to fermented meals and alcoholic drinks.
  • Extra lately, microorganisms have been exploited for the manufacturing of antibiotics, nutritional vitamins and enzymes for use in medication and chemical industries.
  • Moreover, a number of fashionable medication, together with these for most cancers remedy, are pure merchandise or their derivatives.
  • Protists are a nonetheless underexplored supply of pure merchandise probably of curiosity for biotechnological and biomedical purposes.
  • This paper focuses on some examples of bioactive molecules from protists and related micro organism and their potential use in biotechnology.
Key phrases: Bioactive molecules from protists; Climacostol; Chilly-adapted enzymes; Ice binding proteins; Pure merchandise; Secondary metabolites.

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