Recombinant expression and molecular engineering of the keratinase from Brevibacillus parabrevis for dehairing performance.

Keratinase is capable of distinctive degradation of keratin, which provides an eco-friendly approach for keratin waste management towards sustainable development. In this study, the recombinant keratinase (KERBP) from Brevibacillus parabrevis was successfully expressed in Escherichia coli. The purified KERBP had the specific activity of 6005.3 U/mg. It showed remarkable tolerance to various surfactants and also no collagenolytic activity. However, the moderate thermal stability limited its further application. Thus, protein engineering was further adopted to improve its stability. The variants of T218S, S236C and N181D were constructed by site-directed mutagenesis and combinatorial mutagenesis. Compared with the wild type, the t1/2 at 60 °C for the variants T218S, S236C and N181D were 3.05-, 1.18- and 1-fold increase, respectively. Moreover, the double variants N181D-T218S and N181D-S236C significantly improved thermostability with 5.1 and 2.9 °C increase of T50, and prolonging t1/2 at 60 °C with 4.09 and 1.54-fold, respectively. And the catalytic efficiency of the T218S and N181D-T218S variants was also significantly improved. Furthermore, the keratinase displayed favorable ability to dehair wool from skin within 7 h, which showed potential in leather dehairing. Our work contributes to a further insight into the thermostability of keratinase and offers a promising alternative for industrial leather application.

Correlations between growth and wool quality traits of genetically divergent Australian lambs in response to canola or flaxseed oil supplementation.

The correlations between growth and wool traits in response to canola and flaxseed oil supplementation were evaluated in Australian prime lambs. Sixty dual-purpose prime lambs including purebred Merino and crossbred lambs were allocated to one of five treatments of lucerne hay basal diet supplemented with isocaloric and isonitrogenous wheat-based pellets. Treatments were: no oil inclusion (Control); 2.5% canola oil; 5% canola oil; 2.5% flaxseed oil and 5% flaxseed oil, with lamb groups balanced by breed and gender. Each lamb was daily supplemented with 1kg of pellets and had free access to lucerne hay and water throughout the 7-week feeding trial, after a 3-week adaptation. Individual animal basal and supplementary pellet feed intakes were recorded daily, while body conformation traits, body condition scores and liveweights were measured on days 0, 21, 35 and 49. The lambs were dye-banded on the mid-side and shorn before commencing the feeding trial and mid-side wool samples were collected from the same dye-banded area of each lamb at the end of the experiment. Correlations between wool quality traits and lamb performance were non-significant (P>0.05). Oil supplementation had no detrimental effect on lamb growth and wool quality traits (P > 0.05). Gender significantly affected wither height gain and fibre diameter. There were significant interactions between oil supplementation and lamb breed on chest girth. The correlations between clean fleece yield (CFY) and other wool quality traits were moderate ranging from 0.29 to 0.55. Moderate to high correlations between fibre diameter (FD) and other wool quality traits were detected (0.46-0.99) with the strongest relationship between FD and wool spinning fineness (SF). The relationship between CFY and wool comfort factor (CF) were positive, while negative relationships between CFY and the others were observed. A combination of 5% oil supplementation and genetics is an effective and strategic management tool for enhancing feed efficiency and growth performance without negative effects on wool quality in dual-purpose lamb production. This is a good outcome for dual-purpose sheep farmers. It essentially means the absorbed nutrients in supplemented lambs yielded good growth performance without any detrimental impact on wool quality; a win-win case of nutrient partitioning into the synthesis of muscle and wool without compromising either traits.

Dyeing wool and cotton fibres with acidic extract of Hibiscus rosa sinensis flower.

The focus of this work is to extract a natural dye for colouring camel wool as a substitute for synthetic dyes used in the Sadu House of Kuwait. Their target is to keep the tradition of tent and rug production natural in all its manifestations. Therefore, our task was to find an abundant source that provides a colour preferably red to purple. Hibiscus rosa sinensis (HRS) is an abundantly available plant in Kuwait that was explored for extraction of the red dye to colour camel wool permanently. The powdered petals of red flowers of HRS was extracted with 5% acetic acid which yielded a deep red colour that showed a great potential for woollen fibre dyeing. The use of mordants like alum and some metal salts manifested a wide range of fixed colours which intensified at 85 °C. The colours produced had excellent fastness and was accepted by the Sadu House.

Analysis of lecithin treatment effects on the structural transformation of wool fiber using vibrational spectroscopy.

The keratin macromolecule in wool fiber may be found in α-helix or ß-sheet conformations besides a disordered portion. The physical and chemical treatments may cause transformations between α-helix and ß-sheet conformations. The aim of this study was to investigate the influence of lecithin treatment on the wool fiber using the micro-Raman spectroscopy and Fourier transform infrared spectroscopy. Characteristic bands found in the FTIR spectra of wool fibers including the amide A, amide B and amide I-III, which are assigned to the peptide bonds of wool keratin and arise from the amide bonds that link the amino acids. The lecithin treatment didn't affect the peak position of amide bands and only slightly influenced their intensity. It means that the lecithin treatment didn't change the chemical structure of wool fibers. The amide I and III regions, CC skeletal vibration region, and SS bonds vibration regions were analyzed with the Raman microscope. The results indicated the peak area of α-conformation increased gradually by lecithin treatment of the wool fiber, while the peak area of ß-conformation decreased. Therefore, it seems that lecithin treatment of the wool fiber resulted in transformation of ß-sheet to α-helix.

Functionalization of poly(amidoamine) dendrimer-based nano-architectures using a naphthalimide derivative and their fluorescent, dyeing and antimicrobial properties on wool fibers.

Novel naphthalimide-poly(amidoamine) dendrimer fluorescent dyes were synthesized, and their structures were identified and confirmed using different characterization methods such as Fourier transform infrared, (1) H NMR, (13) C NMR, differential scanning calorimetry, elemental analysis and UV-vis spectroscopy. The spectrophotometric studies demonstrated absorption maxima (λmax ) and extinction coefficient (εmax ) values in the ranges of 429-438 nm and 25,635-88,618 L/mol/cm, respectively. The dyeing, fastness and antimicrobial properties of dyed wool fibers were examined. Colorimetric measurements demonstrated a greenish-yellow hue with remarkable fluorescence intensity on dyed wool. Although the fastness properties of naphthalimide dye on wool fibers were poor/moderate, color fastness was appreciably improved through modification of the dye using dendrimers. The results revealed that the newly synthesized dyes are potent antimicrobial agents on wool fibers. Overall, it was deduced that poly(amidoamine) (PAMAM) dendrimers could be exploited as a promising tool in tailoring the different properties of naphthalimide dyes, being suitable for dyeing and antimicrobial finishing agents for wool fibers. Copyright © 2015 John Wiley & Sons, Ltd.

Effect of discarded keratin-based biocomposite hydrogels on the wound healing process in vivo.

Biocompatible keratin-based hydrogels prepared by electron beam irradiation (EBI) were examined in wound healing. As the EBI dose increased to 60 kGy, the tensile strength of the hydrogels increased, while the percentage of elongation of the hydrogels decreased. After 7 days, the dehydrated wool-based hydrogels show the highest mechanical properties (the % elongation of 1341 and the tensile strength of 6030 g/cm(2) at an EBI dose of 30 kGy). Excision wound models were used to evaluate the effects of human hair-based hydrogels and wool-based hydrogels on various phases of healing. On post-wounding days 7 and 14, wounds treated with either human hair-based or wool-based hydrogels were greatly reduced in size compared to wounds that received other treatments, although the hydrocolloid wound dressing-treated wound also showed a pronounced reduction in size compared to an open wound as measured by a histological assay. On the 14th postoperative day, the cellular appearances were similar in the hydrocolloid wound dressing and wool-based hydrogel-treated wounds, and collagen fibers were substituted with fibroblasts and mixed with fibroblasts in the dermis. Furthermore, the wound treated with a human hair-based hydrogel showed almost complete epithelial regeneration, with the maturation of immature connective tissue and hair follicles and formation of a sebaceous gland.

In situ synthesis of nano silver/lecithin on wool: enhancing nanoparticles diffusion.

Silver nanoparticles are being used increasingly in various applications because of their antibacterial properties. It is necessary to lower their direct contact with the skin by embedding in a polymer reducing their side effects. In this study, silver nanoparticles were synthesized inside the wool fibers acted as a polyfunctional ligands. Lecithin as a biological lipid was used to enhance the diffusion of silver ions and nanoparticles into the wool fibers reducing cytotoxicity effects of the nano silver loaded wool. The highest loading efficiency and inhibition zone was observed on the wool with the highest lecithin concentration. Presence of lecithin reduced the rate of nano silver release which results in decreasing the specific coefficient of lethality. Also, the extracted solution of the synthesized silver nanoparticles on the wool has not altered the morphology of L929 fibroblast cells.

Treatment of bleached wool with trans-glutaminases to enhance tensile strength, whiteness, and alkali resistance.

Trans-glutaminases is known as a cross-linking enzyme for proteins. Wool is a proteinous fiber conventionally is treated through several processes to obtain the desirable characteristics. Bleaching is also one of the most important processes usually carried out by using an oxidizing agent in a conventional method. The tensile strength of wool yarns was reduced as a consequence of oxidative bleaching. Here, with the help of microbial trans-glutaminases (m-TGases), a novel bleaching process was disclosed in a way to obtain a bleached wool yarn with no significant reduction in the tensile strength. The results confirmed that the bleached wool yarns with H(2)O(2) could be modified by m-TGases post-treatment. The m-TGases treatment on the bleached wool yarns improved the tensile strength and whiteness along with the higher alkali resistance.

Photo induced silver on nano titanium dioxide as an enhanced antimicrobial agent for wool.

In this study an effective nanocomposite antimicrobial agent for wool fabric was introduced. The silver loaded nano TiO(2) as a nanocomposite was prepared through UV irradiation in an ultrasonic bath. The nanocomposite was stabilized on the wool fabric surface by using citric acid as a friendly cross-linking agent. The treated wool fabrics indicated an antimicrobial activity against both Staphylococcus aureus and Escherichia coli bacteria. Increasing the concentration of Ag/TiO(2) nanocomposite led to an improvement in antibacterial activities of the treated fabrics. Also increasing the amount of citric acid improved the adsorption of Ag/TiO(2) on the wool fabric surface leading to enhance antibacterial activity. The EDS spectrum, SEM images, and XRD patterns was studied to confirm the presence of existence of nanocomposite on the fabric surface. The role of both cross-linking agent and nanocomposite concentrations on the results was investigated using response surface methodology (RSM).

Treatment of wool with laccase and dyeing with madder.

This research has explored the effect of laccase (Denilite II S) on the physical properties of the wool fabric and confirms the anti-felting of wool. In the experiment, laccase was applied to a wool fabric and different characteristics including weight loss, strength, alkali solubility, felting shrinkage, water drop absorption, and dye ability with madder were studied. The surface morphology of the wool fabrics was also observed by scanning electron microscope. The results indicated that the wool fabric treated with laccase has a higher water drop absorption, lower felting shrinkage, and lower values of a* and b*. Treatment of a wool fabric with 10% or lower percentage of laccase reduced the fabric weight but increased the tensile strength. However, using higher concentration of laccase reduced fabric weight and tensile strength. The dyeing of laccase pre-treated wool fabric with madder indicated a lower lightness.

Effects of ultrasound irradiation on wet wool chlorination treatment.

In this study ultrasonic energy is applied for improving wet wool chlorination treatment one of the most well-established methods for reducing wool shrinking related to the felting. Sodium hypochlorite used for providing active chlorine in chlorination bath and samples chlorinated in the presence and absence of ultrasonic irradiation with equal processing parameters. The concentration of active chlorine during the treatment, shrinkage percentage, friction and some mechanical properties of treated samples determined and compared. Scanning Electron Microscope used for investigating the effects of different treatments on removal of wool scales. According to the results it was concluded that applying ultrasonic energy causes desirable mechanical and chemical effects in chlorination process. Cavitation phenomena causes more abrasion on wool surface especially on sharp places i.e., scales and the formation of hydrogen peroxide in ultrasonic bath managing the aggressive reaction of active chlorine on cuticle.

Effects of Psoroptes ovis infection and its control with an ivermectin controlled-release capsule on growing sheep. 2. Evaluation of wool production and leather value.

To evaluate the prophylactic and therapeutic effects of an ivermectin controlled-release capsule (IVM-CRC) on the productivity of growing sheep infested with Psoroptes ovis 24 male and 24 female Merino landrace lambs, 5-6 months old and weighing 21.2-35.0 kg, were used. Sixteen replicates of three animals were formed based on sex and Day 0 body weight. Within each replicate animals were randomly allocated to one of three groups: untreated control; IVM-CRC on Day 0; IVM-CRC on Day 84. For treatment an IVM-CRC for sheep weighing 20-45 kg was used which is designed to deliver ivermectin at a minimum dose of 20 microg/kg/day for 100 days. The lambs were infested with 50-60 P. ovis mites each on days 14 and 21. The animals were shorn on Day 126. After slaughter on Day 127 the quality of the leather produced from their skins was evaluated. The IVM-CRC treatment on Day 0 prevented the establishment of P. ovis. All untreated lambs became infested. The lambs treated with an IVM-CRC on Day 84 became mite-free from Day 112 onwards. The lambs which were treated on Day 0 had significantly (p<0.05) greater clean fleece weight (1.5 kg) and fleece yield (52.1%) than the untreated controls (0.9 kg and 34.1%, respectively) or than lambs treated on Day 84 (1.0 kg and 47.8%, respectively). Clean fleece weight and fleece yield were in the tendency (p=0.055) or significantly (p<0. 001) greater, respectively, for lambs treated on Day 84 than for the untreated controls. The chrome tanned dried crust leather of all untreated controls and six of the 16 lambs treated on Day 84 showed grain surface defects such as discolourations, indurations and coarse, pitted grain. The useful (defect-free) leather size was significantly (p<0.01) higher for the lambs treated on Day 0 (100%) than for the lambs treated on Day 84 (82.7%) and the untreated controls (7.8%). The physical testing revealed that the leather of sheep treated on Day 0 had significantly (p<0.05) higher thickness, elongation break and tear resistance than the leather of both untreated controls and sheep treated on Day 84. There were no significant differences in the physical characteristics of the leather of the untreated controls and the sheep treated on Day 84.

Transsulfuration, protein synthesis rate and follicle mRNA in the skin of young Merino lambs in response to infusions of methionine and serine.

Methionine (Met) is usually the first limiting amino acid for sheep and supplements of Met may increase production of wool and meat. The wool response may be due to an increased supply of cysteine (Cys) from transsulfuration (TS) of Met. Met is catabolized through homocysteine to form Cys when the S from Met is transferred to serine (Ser). We hypothesized that providing additional Met would create a deficiency of Ser and that by simultaneously providing Met and Ser, TS and wool growth could be increased more than by providing Met alone. The effects of i.v. infusions of Met and Ser to young Merino lambs on TS, fractional synthesis rate (FSR) of protein in skin, follicle mRNA and wool growth were examined. Following 4 d of constant i.v. infusion of 3 g Met/d, or 10 g Ser/d or both, the isotope tracers: L-[3-(13)C]Cys, L-[ring-d5]phenylalanine (Phe) and L-[2,3,3-d3]Ser were infused over 8 h to allow for measurements of irreversible loss rate (ILR), and TS in whole body and skin. Skin biopsies were taken for measurement of FSR. Wool growth rate was measured using autoradiography. An infusion of Met significantly (P < 0.05) improved wool growth rate and increased skin FSR, Cys supply from TS and enhanced levels of follicle mRNA (from the K2.10 intermediate filament gene and three gene families encoding keratin associated proteins KAP1, KAP4 and KAP12). The extra Met lowered Ser ILR. The infusion of Ser doubled Ser ILR in the body and increased skin FSR calculated using the Cys tracer in plasma (P < 0.05). However, there were no significant (P > 0.05) changes in TS, skin FSR calculated using the Phe and Ser tracers, follicle mRNA or wool growth rate as a result of Ser infusion. While there were trends towards increased TS and FSR with Ser infusion, the overall lack of significant changes indicates a high capacity for the de novo synthesis of Ser.

The effect of bovine somatotropin treatment on production of lactating angora does with kids.

Fourteen Angora does (35+/-2 kg), each with a single kid and in the first month of lactation, were used to determine ongoing (Period 1) and residual (Period 2) effects of chronic bovine somatotropin (bST) treatment. Specifically, we sought to determine whether chronic bST treatment was capable of improving milk yield, and thus kid growth, and mohair production of nursing does. The experiment consisted of a 2-wk pretreatment period, 5 wk of weekly subcutaneous treatment of slow-release bST (n = 7; Period 1), and a 4-wk posttreatment period (Period 2). The weekly dose of bST was calculated to release 100 microg/(kg BW.d(-1)). To estimate milk production, kids were separated from the does daily for 5 h, and their BW was recorded before and after suckling. The difference in BW was taken as milk production for 5 h. Fiber growth was measured by shearing does at the start of the experiment and at the end of Periods 1 and 2. Dry matter intake and BW of does were not affected by bST (P>.05). Average daily gain of kids that were suckling bST-treated does was higher (P<.05) than for kids of untreated does during Period 1 (184 vs. 139 g/d) but not during Period 2 (140 vs. 136 g/d; P>.10). Treatment with bST did not affect (P>.10) milk composition or clean fleece production in either period. Injection of bST did not affect (P>.10) plasma concentrations of glucose (mean = 49.5 mg/dL), urea N (mean = 19 mg/dL), total protein (mean = 72.5 g/d), or NEFA (mean = 122 microEq/L). During the period of bST treatment, plasma concentrations of somatotropin and IGF-I were increased (P<.05), concentrations of thyroxine and cortisol were decreased (P<.10), and plasma insulin levels were unchanged (P>.10) by bST. In conclusion, treatment of Angora dams with bST did not change DMI or mohair growth, but it improved growth of their kids.

The effects of fibroblast growth factors 1 and 2 on fibre growth of wool follicles in culture.

The effects of fibroblast growth factor-1 (FGF-1) and FGF-2 on fibre growth and follicle function were examined using a previously described procedure to culture wool follicles. Because the FGFs bind to glycosaminoglycan components of the extra-cellular matrix, we also investigated interactions between FGF-1 and FGF-2 with heparin on wool fibre growth. Individual follicles were microdissected from Merino sheepskin and transferred to culture. Follicles increased in length for 6 days, and in all groups, no significant differences in follicle length were observed. Increase in follicle length was associated with fibre growth, follicles maintained normal anagen morphology and incorporated [3H]thymidine into the bulb and outer root sheath cells. Follicles in all treatments continued to produce fibre keratins, as detected by immunohistochemistry. However, the patterns of fibre and cytoskeletal proteins incorporating [35S]methionine in control and treated follicles were significantly different. We found a considerable decrease in the intermediate filament keratins or low sulphur proteins in follicles cultured in the presence of FGF-1 and FGF-2 compared to controls. The majority of proteins detected in these samples were acidic high sulphur proteins. These studies provide evidence for a specific role for the fibroblast growth factors in the regulation of fibre differentiation.

Serum-free culture of wool follicles: effects of nutrients, growth factors and hormones.

A serum-free culture system allowed the continued growth of fibre from follicles for 8-10 days. Fibre growth was responsive to changes in the level of calcium, glucose and amino acids in the culture medium, and was stimulated by the inclusion of insulin (10 micrograms/mL) in the medium. Culture of follicles in the presence of conditioned media from dermal papilla cells or of mitomycin-treated dermal papilla cells had no effect on fibre growth. Neither thyroid hormones nor hydrocortisone altered fibre growth. The progressive decline in fibre growth during follicle culture was accompanied by morphological changes in the follicle bulb. Oxidative damage did not appear to be the cause of these changes as there was no increase in fibre growth rate or longevity when antioxidants were used. This model provides a useful system to study the direct effects of various hormonal, nutritional and growth factors of fibre growth and follicle metabolism.

[Promotion of wool growth and body weight development of sheep by alimentary supplementation with thiocyanate]. / Förderung der Wollbildung und Körpermasseentwicklung beim Schaf durch alimentäre Thiocyanatsupplementierung.

Adult sheep were administered 12 mg thiocyanate (SCN-)/kg body weight orally twice a year in altogether 16 respectively 41 single doses under conventional keeping during the period of stabling (winter) in two consecutive years. Fattening lambs got an analogous SCN- dosage with altogether 4 respectively 34 single doses under intensive keeping. Aided by this temporarily obtained balance of an alimentary SCN- deficiency the following effects were reached during conventional keeping: Significant increase of the wool mass in the greasy state and a tendency to better breading performance. During intensive fattening an increase of the wool mass and of the staple cross-section was proven statistically in the mean for all the test animals, but for the increase of the staple length it could be shown only by comparison of twins.

Effects of epidermal growth factor and transforming growth factor alpha on the function of wool follicles in culture.

The development of a procedure to culture wool follicles from Merino sheep in serum-free conditions has enabled us to investigate the actions of epidermal growth factor (EGF) and transforming growth factor alpha (TGF alpha) on follicle function, including fibre growth. Follicles grown in the absence of growth factors maintained their anagen morphology for 6 days as determined by light microscopy. During this time they incorporated [3H]thymidine into the DNA of the bulb matrix and outer root sheath (ORS) cells and produced fibre keratins as detected by immunohistochemistry. In the presence of EGF and TGF alpha, fibre production ceased after 4 days, as it does following the administration of EGF in vivo. Cessation of fibre growth was not accompanied by regression of the follicle bulb which occurs in vivo. Follicle length growth did not differ significantly from controls and cells in the bulb continued to proliferate. Usually, the structure of the dermal papillae resembled that in control follicles, which was also in marked contrast to changes reported in vivo. In EGF- and TGF alpha-treated follicles, [3H]thymidine continued to be incorporated into DNA of the ORS and bulb after fibre growth ceased. Although wool keratin synthesis ceased, cytokeratins of the epidermis and ORS continued to be produced in the bulb as detected by immunochemistry. These bulb cells were also positive for the periodic acid-Schiff (PAS) reaction indicating the presence of glycogen, a normal component of ORS cells. The observations that cell proliferation continued in the bulb, that glycogen was present and that soft keratins were expressed in these cells suggest that the bulb cell population was induced to differentiate into an ORS phenotype by EGF and TGF alpha.