KR101816622B1 - O-glycosyl hydrolase cel2-KG97 gene from rumen microorganism of black goat and uses thereof - Google Patents
O-glycosyl hydrolase cel2-KG97 gene from rumen microorganism of black goat and uses thereof Download PDFInfo
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Abstract
The present invention relates to the O-glycosyl hydrolase cel2-KG97 gene derived from a black goat rumen microorganism and its use, and more particularly to a novel o-glycosyl hydrolase cel2-KG97 gene selected from a black goat rumen microorganism, Protein products and using the same to feed additives, detergent compositions and methods of making biofuels.
Description
The present invention relates to the O-glycosyl hydrolase cel2-KG97 gene derived from a black goat rumen microorganism and its use, and more particularly to a novel o-glycosyl hydrolase cel2-KG97 gene selected from a black goat rumen microorganism, Protein products and using the same to feed additives, detergent compositions and methods of making biofuels.
The main constituents of plant cell walls are celluloase, hemi-cellulose, and pectin, which are degraded and absorbed by microorganisms through complex and effective processes in herbivore rumen. Rumen microorganisms are extreme anaerobes, including fungi, protozoa and bacteria, and the breakdown of fibrin is mainly responsible for bacteria and fungi. However, most of these microorganisms are not well known because they are difficult to cultivate. Among the ruminal microorganisms known to date, bacteria that are mainly involved in digesting plant cell walls include Fibrobacter succinogenes), luminometer Caucus Playa Pacifica beret Enschede (Ruminococcus flavefaciens), luminometer Cocu live bus (Ruminococcusalbus), Libby brick Booty avoid debris Sol Bence (Butyrivibrio fibrisolvens), oil cake Te Solarium cellulite with a brush Bence (Eubacterium The bacteria belonging to the genus cellulosolvens), frame Beam telra (Prevotella), and some of Clostridium (Clostridium) have been identified. To date, most microbial DNA in the digestive organs of cows, rabbits, and kangaroos, or the metagenome (referred to as the total DNA of environmental microorganisms) has been studied to identify the most abundant enzymes in the rumen, A number of glucanases, beta-glucosidase and cellulosome complexes have been identified through a number of studies.
However, most of the above cases have evolved from soft fungal fibrinogen to food. In industry, degradation ability of fibrinolytic enzyme to harder fibrin source is required.
Black goats evolved to digest feeds of the less coarse fibrinogen, such as twigs, bark, and vegetation. This means that the ruminal microbial population of the black goat evolved differently from other herbivores. Techniques for searching for novel substances using fosmid, cosmid, and bacterial artificial chromosome (GAC) gene banks have been widely used for gene search for promiscuous microorganisms. In this way, a number of fibrinolytic enzyme genes have been found to date.
Under these technical backgrounds, the present inventors have made intensive efforts to accomplish the present invention.
It is an object of the present invention to provide an O-glycosyl hydrolase cel2-KG97 gene derived from a black goat rumen microorganism and its use.
According to one aspect of the present invention, there can be provided an O-glycosyl hydrolase cel2-KG97 gene derived from a black goat rumen microorganism, which comprises the nucleotide sequence shown in SEQ ID NO: 1.
According to another aspect of the present invention, there is provided a
According to another aspect of the present invention, a recombinant vector containing the gene may be provided.
According to another aspect of the present invention, a host cell transformed with the recombinant vector may be provided.
According to another aspect of the present invention, there can be provided a feed additive for promoting fibrin degradation comprising the O-glycosyl hydrolase protein.
According to another aspect of the present invention, a detergent composition comprising the O-glycosyl hydrolase protein can be provided.
According to another aspect of the present invention, there is provided a method for producing a biofuel comprising the step of hydrolyzing a biomass material by adding the O-glycosyl hydrolase protein.
The present invention provides a novel chlorine-derived fibrinolytic enzyme capable of degrading a harder fibrin source. By utilizing the present invention, it is possible to develop a low-cost feed additive against the rise in the price of feed grains, develop biofuels using the most abundant resource on the planet, and apply it to various fields such as a fiber softener and a process for adding a cellulose degrading enzyme can do.
FIG. 1 shows the results of meta genome DNA fragment cleavage conditions and insertion DNA cleavage for the construction of the Fosmid gene bank.
Fig. 2 shows the results of confirming the average insertion size of the phosphide gene bank for the Ad fraction and the Lq fraction.
Fig. 3 shows clonal selection results of inoculation on Esculin-Agar plates and the change of the surrounding brown color according to the esculin degradation.
Figure 4 shows nucleotide blast results according to the present invention.
FIG. 5 shows the major domain identification results according to the protein blast according to the present invention.
Figure 6 shows the major analogous proteins according to the protein blast results of the present invention.
FIG. 7 shows the phylogenetic tree and homology of the highest homologous protein and other fibrinolytic enzymes among the protein blast results of the present invention.
Hereinafter, the present invention will be described in more detail.
The term " superfamily " used in the present invention means a group of proteins having similar amino acid sequences, protein functions, and three-dimensional structures of proteins.
The term " biomass " used in the present invention refers to a whole of a biological organism including a plant carrying out a carbon assimilation process for fixing carbon dioxide using sunlight and a living animal eating it. Various fuels such as alcohol, diesel, and hydrogen produced from the biomass are called biofuel, and biomass such as biomass such as cellulose is proposed as a substitute for chemical substances obtained from petroleum . Plant-based biomass is more advantageous in terms of energy balance than petroleum as raw material in producing various chemical products.
The reason for this is that, in the case of the plant-based biomass, various functional groups including oxygen are included, so that it is possible to convert to a lower activation energy when various chemical products with functional groups are produced.
The term 'biofuel' used in the present invention includes all the byproducts derived from metabolism activities such as feces and living organisms as well as fossil fuels. It is a renewable energy different from fossil fuels, and bioethanol and biodiesel And the like.
The present invention focuses on fibrin degrading ability of a black goat rumen microorganism, and constructs a fosmid gene bank against a black goat rumen microorganism metagenome, from which an exocellulase system, beta-glucosidase activity And the plasmid DNA of the clone was decoded to secure the genetic information. The gene encoding Escherichia coli was transformed into Escherichia coli. The plasmid DNA of the active phosmid clone was extracted using the Qiagen Midi Prep Kit, and the extracted DNA produced 100 times more shotgun sequencing data with an average length of 220 bp. Sequence information was obtained in fasta format. The major domain and similar genes were searched for NCBI blastp sequence for the obtained active fosmid plasmid DNA sequence to identify the fibrinolytic enzyme related gene.
The present invention is very different from the recently disclosed fibrinolytic enzyme gene sequence.
According to one aspect of the present invention, there can be provided an O-glycosyl hydrolase cel2-KG97 gene derived from a black goat rumen microorganism, which comprises the nucleotide sequence shown in SEQ ID NO: 1.
The O-glycosyl hydrolase cel2-KG97 gene according to the present invention may be DNA or RNA encoding an O-glycosyl hydrolase protein. DNA includes cDNA, genomic DNA, or artificial synthetic DNA. The DNA may be single stranded or double stranded. The DNA may be a coding strand or a non-coding strand.
Preferably, the O-glycosyl hydrolase cel2-KG97 gene according to the present invention may comprise the nucleotide sequence shown in SEQ ID NO: 1. Variants of the above base sequences are also included within the scope of the present invention. More specifically, the gene comprises a nucleotide sequence having a sequence homology of 70% or more, preferably 80% or more, more preferably 90% or more, and most preferably 95% or more, from the nucleotide sequence of SEQ ID NO: 1 .
The 'percentage of sequence homology to polynucleotides' is identified by comparing the comparison region with two optimally aligned sequences, and a portion of the polynucleotide sequence in the comparison region is the reference sequence for the optimal alignment of the two sequences (addition or deletion) (I. E., A gap) relative to the < / RTI >
According to an embodiment of the present invention, the O-glycosyl hydrolase cel2-KG97 gene may preferably be derived from a black goat rumen microorganism. However, the present invention is not necessarily limited thereto, and the embodiment of the present invention may be applied to the case where the homology with the O-glycosyl hydrolase cel2-KG97 gene is high (for example, 70% or more, preferably 80% More preferably 90% or more, most preferably 95% or more of the sequence homology) and may be derived from a rumen microorganism of another organism other than a black goat. Methods for sequencing and means for determining sequence homology (e.g., BLAST, etc.) are well known to those skilled in the art.
According to another aspect of the present invention, there is provided a
The range of the O-glycosyl hydrolase protein according to the present invention is a
The term "functional equivalent" means that at least 70% or more, preferably 80% or more, more preferably 90% or more, or more preferably 90% or more, of the amino acid sequence shown in SEQ ID NO: 2 as a result of addition, 2, and most preferably at least 95%, of the amino acid sequence shown in SEQ ID NO: 2, and has substantially the same physiological activity as the protein consisting of the amino acid sequence shown in SEQ ID NO: 2. The term " substantially homogenous bioactivity " means the same O-glycosyl hydrolase activity.
The present invention also includes fragments, derivatives and analogues of O-glycosyl hydrolase proteins. The terms "fragment", "derivative" and "analog" refer to polypeptides having substantially the same biological function or activity as the O-glycosyl hydrolase protein according to the present invention.
Fragments, derivatives and analogs of the O-glycosyl hydrolase protein according to the present invention can be obtained by (i) replacing one or more conservative or non-conservative amino acid residues (preferably conservative amino acid residues) (Ii) a polypeptide having a substituent (s) at one or more amino acid residues, or (iii) another compound that is capable of extending the half-life of the polypeptide (Iv) an additional amino acid sequence (e.g., a sequence derived from a sequence selected from the group consisting of a leader sequence, a secretion sequence, a sequence used to purify the polypeptide, a proteinase A proteinogen sequence or a fusion protein). The fragments, derivatives and analogs defined in accordance with the present invention are well known to those skilled in the art.
The amino acid sequence represented by SEQ ID NO: 2, that is, the polynucleotide encoding the mature polypeptide, is a coding sequence that encodes only the mature polypeptide; Sequences encoding mature polypeptides and various additional coding sequences; Mature polypeptides (and any additional coding sequences) and sequences coding for noncoding sequences.
The term " polynucleotide encoding the polypeptide " refers to a polynucleotide encoding a polypeptide, or a polynucleotide further comprising additional coding and / or noncoding sequences.
The present invention also relates to variants of said polynucleotides encoding polypeptides comprising the same amino acid sequence as described above, or fragments, analogs and derivatives thereof. Polynucleotide variants can be naturally occurring allelic variants or non-naturally occurring variants. The nucleotide mutant includes a substitution mutant, a deletion mutant, and an insertion mutant.
As is known to those of ordinary skill in the art, an allelic variant is an alternative to a polynucleotide, which may comprise one or more substituted, deleted or inserted nucleotides, Lt; RTI ID = 0.0 > polypeptide < / RTI >
According to another aspect of the present invention, a recombinant vector containing the gene may be provided.
The vector according to the invention can typically be constructed as a vector for cloning or expression. In addition, the vector according to the present invention can be constructed by using prokaryotic cells or eukaryotic cells as hosts. For example, when the vector according to the present invention is an expression vector and a prokaryotic cell is used as a host, a strong promoter (for example, pL promoter, trp promoter, lac promoter, T7 promoter, tac promoter, etc.) , Ribosome binding sites for initiation of detoxification, and transcription / translation termination sequences. When E. coli is used as the host cell, the promoter and operator site of the E. coli tryptophan biosynthesis pathway, and the left-handed promoter of the phage (pL promoter) can be used as a regulatory region.
Herein, the vectors that can be used in the present invention include plasmids (e.g., pSC101, ColE1, pBR322, pUC8 / 9, pHC79, pGEX, pQE80L series, pET series and pUC19, etc.) commonly used in the technical field to which the present invention belongs, (E.g., gt4B, -Charon, z1, and M13) or viruses (e.g., SV40, etc.).
In addition, when the vector according to the present invention is an expression vector and the eukaryotic cell is a host, a promoter derived from the genome of a mammalian cell (e.g., a metallothionine promoter) or a mammalian virus (e.g., Adeno Virus late promoter, vaccinia virus 7.5K promoter, SV40 promoter, cytomegalovirus promoter, and tk promoter of HSV) can be used, and generally have a polyadenylation sequence as a transcription termination sequence.
The vectors according to the present invention may be used as selectable markers and include antibiotic resistance genes commonly used in the art to which the present invention belongs, for example, ampicillin, gentamycin, carbenicillin, chloramphenicol, streptomycin, kanamycin, , ≪ / RTI > neomycin, and tetracycline.
According to another aspect of the present invention, a host cell transformed with the recombinant vector may be provided.
The host cell for stably transforming the vector according to the present invention into a prokaryotic cell and then expressing the vector may be a host cell commonly used in the technical field of the present invention, for example, E. coli JM109 , E. coli BL21 , E. coli RR1 , E. coli LE392 , E. coli B , E. coli X 1776 , E. coli W3110, Bacillus subtilis, and Bacillus thuringiensis, or enterobacteria and strains such as Salmonella typhimurium, Serratia marcesensus and various Pseudomonas spp. Can be used.
When the vector according to the present invention is transformed into eukaryotic cells, yeast ( Saccharomyce cerevisiae ), insect cells, human cells (for example, Chinese hamster ovary, W138, BHK, COS-7, 293, HepG2, 3T3, RIN and MDCK cell lines) and plant cells.
The method of delivering a vector according to the present invention into a host cell can be carried out by the CaCl 2 method, Hanahan, D., J. MoI. Biol., 166: 557-580 (1983) ) And electric drilling method. When the host cell is a eukaryotic cell, the vector can be injected into the host cell by microinjection, calcium phosphate precipitation, electroporation, liposome-mediated transfection, DEAE-dextran treatment, gene bombardment, have.
According to another aspect of the present invention, there can be provided a feed additive for promoting fibrin degradation comprising the O-glycosyl hydrolase protein.
Feed additives according to the present invention include organic acids such as citric acid, fumaric acid, adipic acid, lactic acid and malic acid; Phosphates such as sodium phosphate, potassium phosphate, acid pyrophosphate, polyphosphate (polymerized phosphate) and the like; Natural antioxidants such as polyphenol, catechin, alpha-tocopherol, rosemary extract, vitamin C, green tea extract, licorice extract, chitosan, tannic acid, phytic acid and the like have.
The feed additive according to the present invention may include adjuvant components such as amino acids, inorganic salts, vitamins, antibiotics, antimicrobials, antioxidants, antifungal enzymes and other microbial agents in the form of live cells; Grain, for example, ground or crushed wheat, oats, barley, corn and rice; Vegetable protein feedstuffs, for example, based on rapeseed, soybeans and sunflower; Animal protein feeds such as blood, meat, bone meal and fish meal; A dry component consisting of sugar and dairy products, for example various powdered milk and whey powders; Lipids such as animal fat and vegetable fat optionally liquefied by heating; Nutritional supplements, digestion and absorption enhancers, growth promoters, disease prevention agents, and the like.
The feed additive according to the present invention may be in the form of a dry or liquid preparation, and may contain an excipient for feed addition. Examples of the excipient for adding the feed include zeolite, corn or rice bran, and the present invention is not limited to the above examples.
The feed additive according to the present invention may be used as an enzyme preparation, for example, a lipase such as lipase, a phytase which decomposes phytic acid to form phosphate and inositol phosphate, starch and glycogen Amylase, which is an enzyme that hydrolyzes the α-1,4-glycoside bond, phosphatase, which is an enzyme that hydrolyzes the organic phosphate ester, and maltose, And a conversion enzyme which hydrolyzes maltase and saccharose to produce a glucose-fructose mixture, and the like.
The feed additive according to the present invention may be administered to animals alone or in combination with other feed additives in edible carriers. The feed additives can also be administered as top dressing or they can be mixed directly with the livestock feed or separately from the feed, in separate oral formulations, or in combination with other ingredients.
A single daily intake or divided daily intake can be used as is commonly known in the art.
The animal in which the feed additive according to the present invention can be used can be, for example, an animal such as a cow, a cow, a calf, a pig, a pig, a sheep, a goat, a horse, a rabbit, a dog, , Poultry such as ducks, geese, turkeys, quails, small birds, and the like, and are not necessarily limited to the above examples.
The amount of O-glycosyl hydrolase according to the present invention contained in the feed additive according to the present invention is not particularly limited, but as is commonly known in the art to which the present invention pertains, long-term survival in the digestive tract of livestock, Lt; RTI ID = 0.0 > fibrous < / RTI >
According to another aspect of the present invention, a detergent composition comprising the O-glycosyl hydrolase protein can be provided.
The detergent composition according to the present invention may be in the form of a part and a part of an aqueous detergent composition, a non-aqueous liquid detergent composition, a cast solid, a granular form, a granular form, a compressed tablet, a gel, a paste or a slurry. The detergent composition can be used to quickly remove food stains, food residue films, and other small amounts of food compositions.
The detergent compositions according to the present invention may be effective in removing dried stains through the catalytic-mediated hydrolysis of starch polysaccharides.
The detergent composition according to the present invention may be provided in the form of a detergent composition for cleaning hard surfaces, a detergent composition for cleaning fabrics, a detergent composition for washing dishes, a detergent composition for oral cleaning, a detergent for cleaning denture or a contact lens cleaning solution .
According to another aspect of the present invention, there is provided a method for producing a biofuel comprising the step of hydrolyzing a biomass material by adding the O-glycosyl hydrolase protein.
The biomass material according to the present invention is a biomass material containing a high molecular weight carbohydrate. The method for preparing a biofuel according to the present invention comprises: pre-treating a biomass material containing a high molecular weight carbohydrate; Hydrolyzing the pretreated biomass with an enzyme; And a step of fermenting the hydrolyzed biomass material. The method of manufacturing the biofuel may be performed by a method of manufacturing a biofuel commonly known in the art.
In the method for producing a biofuel according to the present invention, the biomass may be selected from the group consisting of starchy (cereals, potatoes), cellulose (herb, wood, chaff) Organic material such as manure, carcass, microbial cells and paper and food waste derived therefrom), and the like, and the present invention is not limited to the above examples.
Hereinafter, the present invention will be described in more detail by way of examples. However, the following examples are illustrative of the present invention and the contents of the present invention are not limited to the following examples.
Example
Isolation and DNA extraction of black goat rumen microorganisms
Two males and one female black goat were raised for only one month with rice straw and mineral supplements, and then slaughtered. The rumen was obtained from the four rumen, and the content was obtained. The obtained contents were first separated into a liquid phase (Lq) and an adherent phase (Ad) by using gauze. Feed scoop Ad was obtained by suspending the residue after filtering with gauze in 0.15% (volume / volume) Tween-80, PBS solution, and recovering the microorganisms attached to rice straw. A total of 5 g of the precipitated sample was prepared and extracted with CTAB (hexadecylmethylammonium bromide) and SDS (sodium dodecyl sulfate).
Black goat rumen microbe fosmid Construction of gene bank
For the DNA extracted from Ad and Lq, the amount of several HindIII restriction enzymes was adjusted for the appropriate truncation conditions to construct a fosmid gene bank. As a result, Ad was determined to use 5 units of HindIII and 2.5 units of Lq (Fig. 1). The partially cleaved DNA fragments were clipped from 10 kb to 50 kb using the Pulse-Field Gel electrophoresis method, from which the insert DNA fragments were again prepared. The prepared DNA was subjected to terminal smoothing reaction and ligated to the pCC1FOS vector by ligation reaction. After the ligation reaction, it was transformed into DH5alpha E. coli by electrical shock method to construct a gene bank. In order to confirm the average insertion size of the constructed fosmid gene bank, NotI enzyme treatment was performed and it was confirmed by electrophoresis that 30 kb for Ad and 31 kb for Lq (FIG. 2). For each of these, 150 plates of 384 well plates were inoculated on cryopreservation media and stored in a cryogenic freezer (-70 ° C).
Esculin ( Esculin ) Decomposing activity Fosmid ( fosmind ) Clone starter
A total of 115,200 clones of Ad and Lq phosphodiester clones were inoculated with 0.5 μl of LB chloramphenicol agar medium containing 1.65% esculin iron substrate in 384 clone units using a VIAFLO 384 instrument from INTEGRA, Time. Clones having the activity of degrading esculin in the cultured plate exhibited a brown color around them, and thus active clones were selected visually (Fig. 3).
Esculin ( Esculin ) Decomposition activity Fosmid Sequence decode for clones
The fosmid plasmid extraction was carried out using the Qiagen plasmid midi prep kit for the phosmid clone Ad_006O06 identified above. The extracted plasmid DNA was averaged 220 bp in length using an Ion Personal Genome Machine (PGM) 100 times more shotgun sequencing data. The resulting data was converted to fasta format and assembled using the Newbler 2.5 de novo assembler program. A total of three consensus sequences were constructed, and a total of 34,870 bp of metagenomic sequence information was obtained with a length of 22,495 bp, 9,103 bp, and 3,272 bp, respectively.
Sequence List
- Sequence Locations: National Livestock Research Institute, RDA
- Black goat derived from the sequence: Rumen obtained from slaughtering after feeding for 3 months (2 males, 1 female) born in the livestock genetics laboratory in the National Livestock Research Institute, RDA Microbial sample
- Fosmid gene bank vector: pCC1Fos
- Genomic Fosmid clone designation: Ad_023A14
- DNA sequence of the gene (SEQ ID NO: 1) Length: 1,467 bp
- the amino acid sequence of the gene (SEQ ID NO: 2) length: 488 amino acids
- DNA and protein sequence of the gene: See the following table and sequence listing
Gene search and similarity survey
The inserted DNA sequence of fosmid clones was searched for the gene region using the MetaGeneMark program. Regarding the genetic information, the sequence similarity information of each gene was firstly obtained by protein blast search of NCBI. In this process, a novel cellulolytic enzyme gene was extracted by referring to the main domain search result and the similar protein. As a result of nucleotide blast (Fig. 4) as of October 2015, Cel2-KG97 can be regarded as a novel gene sequence without a similar sequence to date for the entire 1,467 bp gene. And, as a result of the protein blast search, the
The product of the cel2-KG97 gene, which is a kind of fibrinolytic enzyme O-glycosyl hydrolase according to the present invention, can be used not only as a feed additive but also as a detergent and a bio-fuel, have.
While the present invention has been particularly shown and described with reference to specific embodiments thereof, those skilled in the art will appreciate that such specific embodiments are merely preferred embodiments and that the scope of the present invention is not limited thereby. something to do. It is therefore intended that the scope of the invention be defined by the claims appended hereto and their equivalents.
<110> RURAL DEVELOPMENT ADMINISTRATION <120> O-glycosyl hydrolase cel2-KG97 gene from rumen microorganism of black goat and uses thereof <130> NPF-29354 <160> 2 <170> PatentIn version 3.2 <210> 1 <211> 1467 <212> DNA <213> unidentified bacterium <400> 1 ccctgctcgt gagaagccgg acgagcccgt cgtgaacccg cccgcgccgg tcgacggccc cattgccacc 120 gtgtatacaa cctccgccaa ggaacggttt acggagagca cgctgccgct gggcaagccg 180 gaggacgcgc atttctacaa ggtggagcgc agcggggagg ctttccagga ggtggacggt 240 ttcggcctgg ccatcaccca ggcgagctgc tacaacctgc ggttgatgcc ggaggatgcg 300 cgggacgcct tcctgacgga ggtattcagc cgggaagagg gcctcggctc ctccctgatc 360 cgggtgtgca tcggcggttc cgatttctcc ctggacgagt tcacctggtg tgacgagtcg 420 gggatggaga atttcgccgt ccatccgctg gacgaggaat ggctgttccc ggtcctggac 480 cagatcttcc ggatcaatcc ggcggtgaag atcatcgcct cgccctggag ctgtcccact 540 tggatgaagg tgtccgacag cggcggcttc ttcgaccaat ggaggggcgg ccggctgaat 600 cctgctcatt atcaggaata tgccgattat ttcgtgcgct gggtccggga gatggaaagc 660 cgcggctatc ccatctatgc cgtcaccctg cagaacgaac cgctcaacga gtggaataat 720 tccatgtcgc tgtacatgac ctggcaggag cagcgggatt tcattaagca ggccgtgggg 780 ccggcattcc gcgctgcggg actcaagacc aagatcctgc ttttcgacca caattacaat 840 tacgacgaca tcgcatccca gcaggattat ccgatccgga tcctgcagga cgcggaggcg 900 gcgcagtacg tggccggatc ggcctggcac agctatgggg gaaacgtgtc cgcgctggac 960 aaggtccatg gcgccttccc cgacaaggac atctatttca cggaagcctc catcggctca 1020 tggggttata gctttgacgg ttgtctgatc aatgatttcc gggacatttt cctgggaacg 1080 ctgggtcggt acgggaaagg agtcaccctg tggaacctga tgctggatga cgaggggaaa 1140 ccctatcgcc cgggcgggtg cagcacatgt tttggcgccg tgaccctttc ctcttccgac 1200 cacaggacca ttacccgcaa cagccactat tacaatgtgg cgcactgttc caaggtcttg 1260 ctgcccggcg ctgtgcggtt ggggaccaag ggttacgaaa ccgccggcct cacctaccag 1320 tggtatcgga atcccgacgg ctcccaggcg ttgctgctgc tcaatgaggg gttttctgcc 1380 gccatggtga atttcgtgac gggcaaatat tccatcagtt gtaaggtccc cgccaagtcc 1440 attcagtcca tccgctggga ggagtga 1467 <210> 2 <211> 488 <212> PRT <213> unidentified bacterium <400> 2 Met Lys Met Pro Phe Ser Leu Ile Pro Ala Ala Leu Ala Ala Leu Leu 1 5 10 15 Val Gly Cys Gly Glu Lys Pro Asp Glu Pro Val Val Asn Pro Pro Ala 20 25 30 Pro Val Asp Gly Pro Ile Ala Thr Val Tyr Thr Thr Ser Ala Lys Glu 35 40 45 Arg Phe Thr Glu Ser Thr Leu Pro Leu Gly Lys Pro Glu Asp Ala His 50 55 60 Phe Tyr Lys Val Glu Arg Ser Gly Glu Ala Phe Gln Glu Val Asp Gly 65 70 75 80 Phe Gly Leu Ala Ile Thr Gln Ala Ser Cys Tyr Asn Leu Arg Leu Met 85 90 95 Pro Glu Asp Ala Arg Asp Ala Phe Leu Thr Glu Val Phe Ser Arg Glu 100 105 110 Glu Gly Leu Gly Ser Ser Leu Ile Arg Val Cys Ile Gly Gly Ser Asp 115 120 125 Phe Ser Leu Asp Glu Phe Thr Trp Cys Asp Glu Ser Gly Met Glu Asn 130 135 140 Phe Ala Val His Pro Leu Asp Glu Glu Trp Leu Phe Pro Val Leu Asp 145 150 155 160 Gln Ile Phe Arg Ile Asn Pro Ala Val Lys Ile Ile Ala Ser Pro Trp 165 170 175 Ser Cys Pro Thr Trp Met Lys Val Ser Asp Ser Gly Gly Phe Phe Asp 180 185 190 Gln Trp Arg Gly Gly Arg Leu Asn Pro Ala His Tyr Gln Glu Tyr Ala 195 200 205 Asp Tyr Phe Val Arg Trp Val Arg Glu Met Glu Ser Arg Gly Tyr Pro 210 215 220 Ile Tyr Ala Val Thr Leu Gln Asn Glu Pro Leu Asn Glu Trp Asn Asn 225 230 235 240 Ser Met Ser Leu Tyr Met Thr Trp Gln Glu Gln Arg Asp Phe Ile Lys 245 250 255 Gln Ala Val Gly Pro Ala Phe Arg Ala Ala Gly Leu Lys Thr Lys Ile 260 265 270 Leu Leu Phe Asp His Asn Tyr Asn Tyr Asp Asp Ile Ala Ser Gln Gln 275 280 285 Asp Tyr Pro Ile Arg Ile Leu Gln Asp Ala Glu Ala Ala Gln Tyr Val 290 295 300 Ala Gly Ser Ala Trp His Ser Tyr Gly Gly Asn Val Ser Ala Leu Asp 305 310 315 320 Lys Val His Gly Ala Phe Pro Asp Lys Asp Ile Tyr Phe Thr Glu Ala 325 330 335 Ser Ile Gly Ser Trp Gly Tyr Ser Phe Asp Gly Cys Leu Ile Asn Asp 340 345 350 Phe Arg Asp Ile Phe Leu Gly Thr Leu Gly Arg Tyr Gly Lys Gly Val 355 360 365 Thr Leu Trp Asn Leu Met Leu Asp Asp Glu Gly Lys Pro Tyr Arg Pro 370 375 380 Gly Gly Cys Ser Thr Cys Phe Gly Ala Val Thr Leu Ser Ser Ser Asp 385 390 395 400 His Arg Thr Ile Thr Arg Asn Ser His Tyr Tyr Asn Val Ala His Cys 405 410 415 Ser Lys Val Leu Leu Pro Gly Ala Val Arg Leu Gly Thr Lys Gly Tyr 420 425 430 Glu Thr Ala Gly Leu Thr Tyr Gln Trp Tyr Arg Asn Pro Asp Gly Ser 435 440 445 Gln Ala Leu Leu Leu Leu Asn Glu Gly Phe Ser Ala Ala Met Val Asn 450 455 460 Phe Val Thr Gly Lys Tyr Ser Ile Ser Cys Lys Val Pro Ala Lys Ser 465 470 475 480 Ile Gln Ser Ile Arg Trp Glu Glu 485
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Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR101437491B1 (en) | 2013-06-26 | 2014-09-05 | 대한민국 | xylanase-active enzyme cel10-CBM6-KG61 gene from rumen microorganism of black goat and uses thereof |
| KR101503860B1 (en) | 2013-06-26 | 2015-03-18 | 대한민국 | cellulase cel45-KG80 gene from rumen microorganism of black goat and uses thereof |
| KR101503856B1 (en) | 2013-06-26 | 2015-03-19 | 대한민국 | xylanase-active enzyme cel10-CBM6-KG60 gene from rumen microorganism of black goat and uses thereof |
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Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR101437491B1 (en) | 2013-06-26 | 2014-09-05 | 대한민국 | xylanase-active enzyme cel10-CBM6-KG61 gene from rumen microorganism of black goat and uses thereof |
| KR101503860B1 (en) | 2013-06-26 | 2015-03-18 | 대한민국 | cellulase cel45-KG80 gene from rumen microorganism of black goat and uses thereof |
| KR101503856B1 (en) | 2013-06-26 | 2015-03-19 | 대한민국 | xylanase-active enzyme cel10-CBM6-KG60 gene from rumen microorganism of black goat and uses thereof |
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