Comparison of composition, enzyme activity and selected functional properties of potato proteins isolated from potato juice with two different expanded bed resins

Protein concentrates prepared by expanded bed adsorption (EBA) chromatography from industrial potato juice (PJ) were analysed for chemical composition, color, enzyme activities, thermal properties and selected functional properties (solubility and emulsifying stability). Two EBA multi-modal resins, MIMO I-45 and MIMO 1300 (UpFront Chromatography), were employed under various pH conditions resulting in four potato protein concentrates, A-D. Concentrate B contained an electrophoretically pure protease inhibitor fraction (20-21 kDa), whereas concentrate A, C and D contained both patatin (41 kDa) and protease inhibitors. The potato protein concentrates were explored for the presence of transitions from native to denatured states using differential scanning calorimetry (DSC). Concentrate C had lower heat of transition (ΔH) and T-onset than the other concentrates. The concentrate containing protease inhibitors exhibited the highest denaturation temperature and enthalpy. All concentrates differed significantly (P < 0.05) in color brightness, with concentrate B and D emerging as the brightest. The solubility of the concentrates was evaluated at pH 6 and pH 4.5. All concentrates had lower solubility at pH 4.5 than at pH 6 (70-80%). The stability of emulsions (1% protein, 20% oil, 0.08% xanthan gum) against creaming was analysed with a new method based on the Single electrode Capacitance Probe (SeCaP) technology. Small differences among concentrates were observed by the new SeCaP method.     

SZ蛋白酶对羊毛的减量和防毡缩性能的影响

在用蛋白酶对羊毛织物进行改性处理的研究中，获得一种对羊毛具有独特处理效果的蛋白酶ＳＺ。本文主要就ＳＺ蛋白酶对羊毛的减量特性及其防毡性能的影响进行研究。ＳＺ蛋白酶采用Ｐｅｒｚｙｍ工艺对羊毛进行减量处理，可有效地防止羊毛毡缩。减量处理对羊毛的损伤较小，实际应用可以被接受。研究表明，羊毛的减量防毡缩处理并不需要完全去除鳞片，对于有效的减量，去除２／３的鳞片可获得最佳的减量处理效果。     

Use of commercial protease preparations to reduce protein and lipid content of maize starch

A method was developed to produce pure maize starch from maize flour using a protease processing step, and additional salt washing and sulphite steeping steps. A range of commercial protease enzymes were evaluated for this purpose. The laboratory scale procedure that was developed, using one protease in particular (Promod P25P, thermolysin), produced relatively pure starch (<0.45% protein). Using the same procedure, but applying to starches which had been produced in advance using traditional wet milling, starch protein contents could be reduced further by 25–50% with the lipid content reduced by up to 25%. The amount of starch damage was minimal using this approach (<1%). This procedure could be developed industrially for a ‘greener approach’ to starch extraction – although it may still be necessary to incorporate sulphite steeping stages to facilitate protein solubilisation and extraction.     

Stepwise optimisation of enzyme production in solid state fermentation of waste bread pieces

When it is not consumed, bread presents a major source of food waste, both in terms of the amount and its economic value. However, bread also possesses the characteristics of an ideal substrate for solid state fermentation. Yet nearly all wasted bread ends up in landfill sites, where it is converted into methane by anaerobic digestion. Governments are finally taking action and, according to the EU Landfill Directive, for example, biodegradable municipal waste disposed into landfills must be decreased to 35% of 1995 levels, by 2020. Solid state fermentation of waste bread for the production of value added products is a novel idea, which could help with the achievement of this target. In this study, glucoamylase and protease production from waste bread pieces, via solid state fermentation, was investigated in detail. The optimum fermentation conditions for enzyme production were evaluated as, 20 mm particle size, 1.8 (w/w, db) initial moisture ratio, and duration of 144 h. Under these conditions, glucoamylase and protease activities reached up to 114.0 and 83.2 U/g bread (db), respectively. This study confirms that waste bread could be successfully utilised as a primary raw material in cereal based biorefineries.     

Purification and characterisation of a novel protease from Cordyceps sinensis and determination of the cleavage site motifs using oriented peptide library mixtures

A novel protease, from the edible fungus Cordyceps sinensis, was purified and characterised. Its cleavage site motifs were determined by oriented peptide library mixtures and validated by synthetic peptides and natural proteins.     

Protein hydrolysis by protease isolated from tuna spleen by membrane filtration: A comparative study with commercial proteases

Membrane filtration is considered to be a low-cost and large scale method for recovery and purification of enzymes. The trypsin-like serine protease (TSP) was recovered and purified by microfiltration and ultrafiltration from the extract of the spleen of the yellowfin tuna (Thunnus albacores). The partially purified TSP has the activity of 96.5 U/mL and purity of 74.2 U/mg protein based on casein digesting unit. It has the molecular weight of 24 kDa proved by SDS-PAGE. The potential application of TSP in the protein hydrolysis was investigated in comparison to the use of two commercial proteases, i.e. Alcalase^® 0.6 L and Delvo-Pro under their optimal hydrolysis conditions. The effects of enzymes, substrates and enzyme to substrate ratio on the degree of hydrolysis (DH) were studied. Alcalase showed the highest DH for both casein and soybean isolate. TSP showed higher DH than Delvo-Pro when casein was employed as the substrate. However, TSP showed the lowest DH when soybean protein was used as the substrate. The present study proved that TSP recovered and purified from the tuna canning waste by membrane filtration can be used for protein hydrolysis as well as other commercial proteases.     

Preparation of a rice bran enzymatic extract with potential use as functional food

The production, stabilization, by enzymatic treatment, physicochemical composition, and biological properties (including the anti-proliferative activity), of a water-soluble rice bran enzymatic extract (RBEE) are described. The main component of RBEE is proteins (38.1%) – in the form of peptide and free amino acids – having a 6% content of sulfur amino acids. The second component is fat (30.0%), with oleic and linoleic acids as the major components, and 1.2 mg/g of γ-oryzanol. Carbohydrates (14.2%) are comprised mainly of slowly absorbed carbohydrates. Preliminary studies on the anti-proliferative effect of RBEE on leukemia tumor cell growth in vitro are also reported. This property makes RBEE potentially useful as a functional food for the treatment and prevention of chronic pathological states associated with abnormal proliferation of cells, as is the case with cancer.     

N-linked glycosylation of proteinase B precursors of the yeast Saccharomyces cerevisiae is not required for proper targeting or processing of the enzyme.

Proteinase B precursors are modified by an N-linked carbohydrate side chain at Asn 314. Glycosylation at this position is not required for proper localization, processing, or activation of the enzyme.     

Bowman-Birk inhibitors in lentil: Heterologous expression,functional characterisation and anti-proliferative properties in human colon cancer cells

A full-length cDNA, encoding a Bowman-Birk protease inhibitor (BBI), was isolated from lentil immature seeds. The deduced amino acid sequence was longer than that of the BBI extracted from lentil seeds and contained two binding sites; the first inhibitory site inhibits trypsin whereas the second one inhibits chymotrypsin. In order to characterize this lentil BBI, a longer (complete) and its C-terminally processed (mature) form were heterologously expressed in the yeast Pichia pastoris. The recombinant BBI proteins proved to be active against trypsin and chymotrypsin, showing K_i values at nanomolar levels. Mass spectrometry analysis revealed that complete BBI was composed of an array of molecular masses, whereas mature BBI showed the presence of a major peak of the expected size. The effects of mature BBI on the growth of human colon adenocarcinoma HT29 and colonic fibroblast CCD-18Co cells were evaluated. Lentil BBI was able to inhibit the growth of such cells at concentrations higher than 19 μM, in a concentration-dependent manner; by contrast, the CCD18-Co cells were unaffected. These data broaden our knowledge of the beneficial biological activities of naturally-occurring BBI proteins and address the need for systematic evaluation of natural variants in order to design novel strategies in preventive medicine.     

Engineering of protease-resistant phytase from Penicillium sp.: High thermal stability, low optimal temperature and pH

A new and simple method based on the mechanism of detoxification of metallothionein was developed by using a water-soluble porphyrin and Zn(II)-bound metallothionein for evaluating heavy metal toxicity. Labile Zn(II) ions were released when toxic metal ions such as Cu(II), Pb(II), Bi(III), Cd(II), Hg(II), Co(II), Ag(I), and Ni(II) bound to Zn(II)-bound metallothionein. The water-soluble porphyrin, 5,10,15,20-tetraphenyl-21H,23H-porphinetetrasulfonic acid, a chromogenic reagent that is highly sensitive to Zn(II), formed a complex with the labile Zn(II) ions. The absorption change at 423 nm resulting from the formation of the Zn(II)–porphyrin complex was used to evaluate the toxicity of sample solutions containing different metal ions. The absorption change was well correlated with the toxicity, which was evaluated by a bioluminescence inhibition assay using the bioluminescent bacteria Vibrio fischeri. This observation indicated that the absorption change determined by our method was a good indicator of heavy metal toxicity. The proposed method was more sensitive than conventional bioassays and could be used to detect metal toxicity at submicromolar concentrations of toxic metal ions.