A Biofinishing System

Abstract

The present disclosure relates to a biofinishing system comprising a combination of cellulases, in particular a biofinishing system comprising a combination of GH45 cellulases. The present disclosure further relates to a process for treating a cellulose-containing textile comprising biofinishing the cellulose-containing textile with a combination of GH45 cellulases.

Claims

1. An enzyme composition comprising, a first polypeptide having GH45 cellulase activity and biofinishing activity, and a second polypeptide having GH45 cellulase activity and biofinishing activity.

2. The enzyme composition of claim 1, wherein the first polypeptide and the second polypeptide exhibits synergy in an assay that measures biofinishing activity.

3. The enzyme composition of claim 1, wherein the first polypeptide and the second polypeptide are endoglucanases.

4. The enzyme composition of claim 1, wherein the first polypeptide is in an amount of from about 30% to about 330%, by weight of the second polypeptide.

5. The enzyme composition of claim 1, wherein the first polypeptide or the second polypeptide has at least 80% sequence identity to SEQ ID NO: 2, the mature polypeptide of SEQ ID NO: 4, the mature polypeptide of SEQ ID NO: 6 or the mature polypeptide of SEQ ID NO: 8.

6. The enzyme composition of claim 5, wherein the first polypeptide or the second polypeptide comprises or consists of SEQ ID NO: 2, the mature polypeptide of SEQ ID NO: 4, the mature polypeptide of SEQ ID NO: 6 or the mature polypeptide of SEQ ID NO: 8.

7. A nucleic acid construct or expression vector comprising a first polynucleotide encoding a first polypeptide having GH45 cellulase activity and biofinishing activity, and a second polynucleotide encoding a second polypeptide having GH45 cellulase activity and biofinishing activity, operably linked to one or more control sequences that direct the production of the polypeptides in an expression host.

8. A recombinant host cell comprising the nucleic acid construct or expression vector of claim 7.

9. A process for biofinishing a cellulose-containing textile, comprising contacting the cellulose-containing textile with the composition of claim 1 or the recombinant host cell of claim 8.

10. The process of claim 9, wherein the cellulose-containing textile is a cotton-containing textile or man-made cellulose-containing textile.

11. A process for biofinishing a cellulose-containing textile, comprising (a) treating the cellulose-containing textile with a first polypeptide having GH45 cellulase activity and biofinishing activity; and (b) treating the cellulose-containing textile with a second polypeptide having GH45 cellulase activity and biofinishing activity.

12. The process of claim 11, wherein the first polypeptide or the second polypeptide has at least 80% sequence identity to SEQ ID NO: 2, the mature polypeptide of SEQ ID NO: 4, the mature polypeptide of SEQ ID NO: 6 or the mature polypeptide of SEQ ID NO: 8.

13. A process for treating a cellulose-containing textile, comprising (a) desizing; (b) color modification; wherein a first polypeptide having GH45 cellulase activity and biofinishing activity and a second polypeptide having GH45 cellulase activity and biofinishing activity are added before, during or after step (a) and step (b).

14. A process for treating a cellulose-containing textile, comprising (a) desizing; (b) scouring; (c) bleaching; (d) dyeing; wherein a first polypeptide having GH45 cellulase activity and biofinishing activity and a second polypeptide having GH45 cellulase activity and biofinishing activity are added before, during or after step (a), (b), (c) or (d).

15. (canceled)

16. A method of producing a composition comprising a first polypeptide having GH45 cellulase activity and a second polypeptide having GH45 cellulase activity, comprising: (a) cultivating a recombinant host cell of claim 8 under conditions conducive for production of the polypeptides; and (b) recovering the composition.

Description

EXAMPLES

[0245] Materials

[0246] Chemicals used as buffers and substrates were commercial products of at least reagent grade.

[0247] Media

[0248] 1) pH 5.0 buffer with 50 mM acetate: 2.873 g sodium acetate and 0.901 g acetic acid dissolved in 1 L de-ionized water;

[0249] 2) pH 6.5 buffer with 50 mM phosphate: 5.642 g disodium hydrogen phosphate dodecahydrate (Na.sub.2HPO.sub.4.12H.sub.2O) and 5.344 g sodium dihydrogen phosphate dehydrate(NaH.sub.2PO.sub.4.2H.sub.2O) dissolved in 1 L de-ionized water;

[0250] 3) pH 7.5 buffer with 50 mM phosphate:15.045 g disodium hydrogen phosphate dodecahydrate (Na.sub.2HPO.sub.4.12H.sub.2O) and 1.248 g sodium dihydrogen phosphate dehydrate(NaH.sub.2PO.sub.4.2H.sub.2O) dissolved in 1 L de-ionized water;

[0251] 4) pH 8.5 buffer with 50 mM phosphate: 17.607 g disodium hydrogen phosphate dodecahydrate (Na.sub.2HPO.sub.4.12H.sub.2O) and 0.116 g potassium dihydrogen phosphate (KH.sub.2PO.sub.4) dissolved in 1 L de-ionized water.

[0252] Enzymes

[0253] GH45-1: a Staphylotrichum coccosporum GH45 cellulase disclosed as mature polypeptide of SEQ ID NO: 3 in WO2005/054475, and as SEQ ID NO: 2 herein.

[0254] GH45-2: a Thielavia terrestris GH45 cellulase disclosed as the mature polypeptide of SEQ ID NO: 4 in WO 2012/089024 and as the mature polypeptide of SEQ ID NO: 4 herein .

[0255] GH45-3: a Sordaria fimicola GH45 cellulase disclosed as the mature polypeptide of SEQ ID NO: 2 in WO 2014/026630 and as the mature polypeptide of SEQ ID NO: 6 herein.

[0256] GH45-4: Neurospora tetraspermaGH45 cellulase disclosed in the mature polypeptide of SEQ ID NO: 2 in WO 2015/058700 and as the mature polypeptide of SEQ ID NO: 8 herein.

[0257] Fabrics

[0258] Cotton interlock: 40S, bleached, HM-A0008, available from HM Cotton, Guangzhou, China.

[0259] Method

[0260] Weight Loss Determination

[0261] The swatches were placed in a conditioned room (65%+/5% humidity, 20+/1 C.) for 24 hours before they were numbered, weighed by an analytical balance (for samples below 100 g) or a precision balance (for samples over 100 g) and recorded. After treatment, all samples were tumbled dried for 1 hour and conditioned for 24 hours in the conditioned room mentioned as above. For each sample, the weight loss was defined as below:

[00001]$\mathrm{Weight}.Math..Math.\mathrm{loss}.Math..Math.\%=\frac{\left(\mathrm{weight}.Math..Math.\mathrm{before}.Math..Math.\mathrm{treatment}-\mathrm{weight}.Math..Math.\mathrm{after}.Math..Math.\mathrm{treatment}\right)*100}{\mathrm{weight}.Math..Math.\mathrm{before}.Math..Math.\mathrm{treatment}}$

[0262] Pilling Notes Test

[0263] Fabrics including treated and untreated were pre-conditioned in norm climate (65% humidity, 21 C.) for at least 24 hours and then tested for the pilling notes with Nu-Martindale Tester (James H. Heal Co. Ltd, England), with untreated fabrics of the same type as the abraded fabrics on the bottom. A standard pilling test (Swiss Norm (SN) 198525) was carried out after 2000 Revolutions by marking from 1-5, with the meaning defined as below, where 1 shows poor anti-pilling and 5 shows excellent anti-pilling property. Thus the higher the Martindale pilling notes score the more effective the cellulose biopolishing treatment.

[0264] Note 5: No pilling

[0265] Note 4: Slight Pilling

[0266] Note 3: Moderate Pilling

[0267] Note 2: Distinct Pilling

[0268] Note 1: Heavy Pilling

[0269] , notes are allowed

[0270] To make the test result more reliable, 3 separate readings were carried out by different persons for each sample, and the average of the 3 readings was adopted as the final result of pilling notes.

[0271] Protein Content

[0272] The enzyme protein in an enzyme product can be measured with BCA Protein Assay Kit (product number 23225, commercial available from Thermo Fisher Scientific Inc.) according to the product manual.

Example 1

Biopolishing With GH45-1 and GH45-2 in Launder-O-Meter

[0273] Cellulases GH45-1 and GH45-2 were tested on their own and in blending for biopolishing performance in Launder-O-meter (LOM).

[0274] Cotton fabric swatches were cut into about 16 cm*16 cm (about 5 grams each). The swatches were placed in a conditioned room (65% humidity, 21 C.) for 24 hours before they were numbered, weighed by an analytical balance and recorded. The biopolishing was conducted with a LOM. Two conditioned swatches were put into each beaker. 20 big steel balls (total eight of 220 grams) or 4 rubber balls (total weight of 5 grams) were placed in each beaker to supply high and low level of mechanical actions, respectively. The beaker was filled with enzymes according to Table 1 and buffers prepared as described in media part to a total volume of around 100 ml, which could get a liquid to fabric ratio of about 10:1 (v/w).

[0275] The LOM machine was started after the required program was chosen, and it would hold when the temperature reached the pre-set temperature, e.g. 55 C. Each beaker was fitted with a lid lined with 2 neoprin gaskets and closed tightly with a metal clamping device. The beakers were loaded into the preheated LOM. Metal racks were used to accommodate and secure 5 beakers, in the vertical position, in each of the 4 drum positions. The LOM lid was closed and the washing program was continued and the timing was initiated. 1 hour later, all beakers were removed from LOM and the denim samples were transferred to an inactivation solution (2 g/L sodium carbonate) at 80 C. for 10 minutes. Then the swatches were rinsed in hot water for 2 times and in cold water for 2 times and they were tumble-dried (AEG, LAVATHERM 37700, Germany) for 1 hour, conditioned for 24 hours at 65% relative humidity, 21 C. prior to evaluation in weight loss and pilling notes.

[0276] As summarized in Table 1, it is evident that the blendings of GH45-1 and GH45-2 can deliver stronger biopolishing performance than any one of these two on their own, i.e. the two GH45 can work in a synergetic way: at high mechanical aids with 20 steel balls, when applied alone, 0.049 mg/g GH45-1 and 0.038 mg/g GH45-2 as 100% on their own delivered 3.8 and 3.6 in pilling notes, respectively; while when these two products were applied together at the ratio 50:50 or 25:75, the blendings can delivered pilling notes 4.5, which was 0.7-0.9 higher than they were applied on their own; at low mechanical aids with 4 rubber balls, when applied alone, 0.049 mg/g GH45-1 and 0.076 mg/g GH45-2 as 100% on their own delivered 1.5 and 3.5 in pilling notes, respectively, while when these two products were applied together at the ratio 95:5, 86:14, 67:33 and 40:60, the blendings can delivered pilling notes 3.3, 3.6, 3.5, 3.9, which were much higher than GH45-1 alone and similar or stronger than GH45-2 alone. To obtain a same benefits in pilling notes, the risk in weight loss was also obviously reduced with the blendings with different ratios.

Example 2

Biopolishing With GH45-2 and GH45-3 in Launder-O-Meter

[0277] Cellulases GH45-2 and GH45-3 were tested on their own and in blending for biopolishing performance in LOM with the same protocol and procedure as in Example 1 except that the biopolishing trials were conducted at 45 C. and 55 C., respectively.

[0278] From the results as summarized in Table 2, it can be seen that GH45-2 and GH45-3 also had synergetic effects when they were applied together. For example, at 45 C., when applied alone, 0.146 mg/g GH45-2 and 0.115 mg/g GH45-3 as 100% on their own delivered 4.0 in pilling notes, respectively; while when these two products were applied together at the ratio 80:20 or 60:40, the blendings can delivered pilling notes 4.4, which was 0.4 higher than they were applied on their own. At 55 C. similar synergetic effects between these two GH45s were observed. Meanwhile the blendings of GH45-2 and GH45-3 delivered more consistent performance when temperature changed from 45 C. to 55 C.

Example 3

Biopolishing With GH45-1 and GH45-3 in Launder-O-Meter

[0279] Cellulase GH45-1 and GH45-3 were tested on their own and in blending for biopolishing performance in LOM with the same protocol and procedure as in Example 1 except that the biopolishing trials were conducted at 35 C.

[0280] As summarized in Table 3, at 35 C., 0.114 mg/g GH45-1 and 0.0805 mg/g GH45-3, were set as 100% base for blending, respectively. GH45-1/GH45-3 blending with all the blending ratios (90:10, 75:26, 50:50, 25:76, 10:90 for GH45-1/GH45-3) delivered better anti-pilling performance than GH45-1 alone or GH45-3 alone.

Example 4

Biopolishing With GH45-2 and GH45-4 in Launder-O-Meter

[0281] Similar to Example 3, biopolishing trials were conducted to compare cellulases GH45-2 and GH45-4 on their own and in blending.

[0282] As summarized in Table 4, at 55 C., 0.0394 mg/g GH45-2 and 0.0378 mg/g GH45-4 as 100% on their own delivered pilling notes at 3.8 and 2.9, respectively, while the blendings of GH45-2 and GH45-4 at different ratios (30:70, 52:50, 70:30 for GH45-2/GH45-4) can deliver similar performance to GH45-2 and much better performance than GH45-4 alone, which also suggested that these two GH45s can work in a synergetic way.

Example 5

Biopolishing With GH45-1 and GH45-4 in Launder-O-Meter

[0283] Similar to Example 4, biopolishing trials were conducted to compare cellulases GH45-1 and GH45-4 on their own and in blending.

[0284] As summarized in Table 5, at 35 C., 0.114 mg/g GH45-1 and 0.0945 mg/g GH45-4 as 100% on their own delivered pilling notes at 3.1 and 4.3, respectively. When 25%, 50%, 75% and 90% GH45-1 was replaced with the corresponding percentages of GH45-4, the resulting blendings delivered pilling notes in the range of 4.1 to 4.3, which indicated the synergetic effects between these two cellulases.

Example 6

Biopolishing With GH45-2 and GH45-3 in a Jet Dyer

[0285] Celluases GH45-2 and GH45-3 which had been tested in Example 2, were also tested in a production machine in a textile mill.

[0286] In each trial, about 20 kg 100% cotton fabrics which had been scoured and bleached by the mill were loaded into a dyeing machine (Shuangxi ECO Jet). The winch speed and pump pressure were adjusted to make sure the fabric circulate once every 55 seconds and ran smoothly in the machine; 200 L water was loaded in each step to keep the liquor ratio of about 10:1 (water volume/fabric weight). In each trial, the fabric was treated as followed:

[0287] 1) Pre-washed with water at room temperature for 10 min;

[0288] 2) Drained;

[0289] 3) Main-washed with a cellulase bath at 35 C. or 55 C. pH 6-6.5 adjusted with acetic acid, initiated timing when cellulase was loaded into the bath and collected the fabrics after 45, 60, 75, 90 and 105 min;

[0290] 4) Drained;

[0291] 5) Rinsed with water twice;

[0292] 6) Centrifuged and dried.

[0293] The fabrics collected in the process were sent for further evaluation for fuzz level and pilling notes (after Nu-Martindale treatment).

[0294] From Table 6 it can be seen that the two cellulases works in a synergetic way: 0.244 mg/g GH45-2 at 55 C. delivered similar biopolishing performance as 0.380 mg/g GH45-3 at 35 C., while the blending of these two cellulases at 50:50 or 70:30 for GH45-2/GH45-3 at 55 C. delivered a stronger and faster biopolishing than both on their own. In 60 min treatment, GH45-2 and GH45-3 delivered pilling notes 2.0 and 2.5, respectively; while the blendings at two ratios delivered 3.5 and 2.8, respectively. It can also been seen that a similar trend in fuzz level: the blendings delivered an equivalent fuzz level with 15-30 min shorter time than GH45-2 or GH45-3 on their own. It can further been seen that the blending with 50% GH45-2 and 50% GH45-3 delivered a stable biopolishing performance when the main washes were conducted at 35 C. or 55 C.

[0295] The invention described and claimed herein is not to be limited in scope by the specific aspects herein disclosed, since these aspects are intended as illustrations of several aspects of the invention. Any equivalent aspects are intended to be within the scope of this invention. Indeed, various modifications of the invention in addition to those shown and described herein will become apparent to those skilled in the art from the foregoing description. Such modifications are also intended to fall within the scope of the appended claims. In the case of conflict, the present disclosure including definitions will control.