# US 10,308,967 B2 · Milling process

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- Number: US 10,308,967 B2
- Document ID: US10308967B2
- Status: Granted
- Kind: B2
- Application number: 15/033,751
- Filed: 2014-11-26
- Published: 2016-09-08
- Granted: 2019-06-04
- Earliest claimed: 2013-11-26
- Assignees: Novozymes A/S
- Applicants: Novozymes A/S
- Inventors: Zhen Long; Wanghui Xu; Wang Han; Scott R. McLaughlin; Randall Deinhammer; Paria Saunders; Bernardo Vidal, Jr.; Xinyu Shen; Michael John Akerman; Tom Gibbons
- Web page: https://patentgrep.com/patent/US10308967B2

## CPC

- C12P 19/04: Polysaccharides, i.e. compounds containing more than five saccharide radicals attached to each other by glycosidic bonds
- C08B 30/02: Preparatory treatment, e.g. crushing of raw materials {or steeping process (machines for preliminary washing A23N)}
- C08B 30/042: {from cereals or grains}
- C08B 30/044: {from corn or maize}
- C08B 30/046: {from wheat}
- C12N 9/58: derived from fungi
- C12N 9/6408: {Serine endopeptidases (3.4.21)}
- C12P: FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE {(brewing of beer C12C; producing vinegar C12J; producing specific peptides or proteins C07K; producing enzymes C12N9/00; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification C12N15/00; measuring or testing processes involving enzymes or microorganisms C12Q; measuring or testing processes involving nucleic acid amplification reactions C12Q1/6844; fermentation processes to form a food composition, A21 or A23; compounds in general, see the relevant compound class, e.g. C01, C07)}
- C08B: POLYSACCHARIDES; DERIVATIVES THEREOF (polysaccharides containing less than six saccharide radicals attached to each other by glycosidic linkages C07H; fermentation or enzyme-using processes C12P19/00; sugar industry C13; production of cellulose D21)
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)

## Abstract

Provided are processes for treating crop kernels which comprising the steps of a) soaking kernels in water to produce soaked kernels; b) grinding the soaked kernels; c) treating the soaked kernels in the presence of a polypeptide having protease activity.

## Claims

### Claim 1

A process for treating crop kernels, comprising the steps of:
a) soaking kernels to produce soaked kernels;
b) grinding the soaked kernels;
c) treating the soaked kernels in the presence of a polypeptide having protease activity,
wherein step c) is performed before, simultaneously with or after step b),
and said polypeptide is selected from the group consisting of:
(a) a polypeptide having at least 90% sequence identity to the polypeptide of SEQ ID NO: 5, SEQ ID NO: 6, or the mature polypeptide of SEQ ID NO: 2 or SEQ ID NO: 4; and
(b) a polypeptide encoded by a polynucleotide having at least 90% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 1 or SEQ ID NO: 3.

### Claim 2 (depends on 1)

The process of claim 1, wherein the said polypeptide is a polypeptide having at least 95% sequence identity to the polypeptide of SEQ ID NO: 5, SEQ ID NO: 6, or the mature polypeptide of SEQ ID NO: 2 or SEQ ID NO: 4.

### Claim 3 (depends on 1)

The process of claim 1, wherein the polypeptide is a polypeptide encoded by a polynucleotide having at least 95% sequence identity to the mature polypeptide coding sequence of SEQ ID NO: 1 or SEQ ID NO: 3.

### Claim 4 (depends on 1)

The process of claim 1, wherein the polypeptide having protease activity comprises or consists of SEQ ID NO: 2, SEQ ID NO: 4, SEQ ID NO: 5 or SEQ ID NO: 6.

### Claim 5 (depends on 1)

The process of claim 1, wherein the polypeptide having protease activity is from the genus Meripilus, genus Dichomitus, or genus Trametes.

### Claim 6 (depends on 1)

The process of claim 1, further comprising treating the soaked kernels in the presence of a beta-xylosidase.

### Claim 7 (depends on 1)

The process of claim 1, further comprising treating the soaked kernels in the presence of a cellulase and/or a hemicelluase.

### Claim 8 (depends on 1)

The process of claim 1, further comprising treating the soaked kernels in the presence of a xylanase.

### Claim 9 (depends on 1)

The process of claim 1, further comprising treating the soaked kernels in the presence of a cellulolytic composition comprising: 1) a cellulase or a hemicellulase, and 2) a GH61 polypeptide.

### Claim 10 (depends on 1)

The process of claim 1, further comprising treating the soaked kernels in the presence of an acetylxylan esterase.

### Claim 11 (depends on 1)

The process of claim 1, further comprising treating the soaked kernels in the presence of an enzyme selected from the group consisting of an endoglucanase, a xylanase, a cellobiohydrolase I, a cellobiohydrolase II, a GH61 polypeptide, and combinations thereof.

### Claim 12 (depends on 1)

The process of claim 1, wherein said polypeptide is present in an amount of 0.0005 to 1.5 mg enzyme protein per g DS kernels.

### Claim 13 (depends on 1)

The process of claim 1, wherein the crop kernels are from corn, maize, rice, barley, sorghum, bean, fruit hulls, or wheat.

### Claim 14 (depends on 12)

The process of claim 12, wherein said polypeptide is present in an amount of 0.001 to 1 mg enzyme protein per g DS kernels.

### Claim 15 (depends on 12)

The process of claim 12, wherein said polypeptide is present in an amount of 0.01 to 0.5 mg enzyme protein per g DS kernels.

### Claim 16 (depends on 1)

The process of claim 1, wherein said polypeptide is present in an amount of 0.025 to 0.25 mg enzyme protein per g DS kernels.