Microorganism with improved L-alanine-producing ability and method for producing L-alanine using the same

Abstract

The present invention relates to a Corynebacterium mutant strain that has improved L-alanine-producing ability compared to the parent strain due to mutation induction by irradiation, a method for producing the strain, and a method for producing L-alanine using the strain. Since the Corynebacterium mutant strain has improved L-alanine-producing ability compared to the parent strain, L-alanine can be produced with high efficiency and high yield by using the strain.

Description

[Technical Field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims the benefit of priority based on Korean Patent Application No. 10-2022-019075, filed December 30, 2022, and all contents disclosed in the documents of this Korean patent application are incorporated herein by reference.

[0002] The present invention relates to a microorganism with improved L-alanine-producing ability and a method for producing L-alanine using the same. [Background technology]

[0003] L-alanine is an important amino acid with a wide range of applications in the fields of chemistry, food, and medicine. It is an amino acid that imparts a sweet taste and is particularly used in the food industry as a flavor enhancer and nutritional supplement. As of 2020, the global alanine market was valued at over USD 250 million, with an annual production of approximately 500 tons (The Expresswire, 2020; Wendisch, 2014).

[0004] L-alanine is generally obtained through chemical synthesis or enzymatic conversion. Chemical synthesis is achieved by modifying the Strecker synthesis or Bucherer-Bergs method (Legnani et al., 2021), while enzymatic conversion is achieved by using ammonium fumarate from L-aspartate (Takamatsu et al., 1982). Most industrial production is achieved through enzymatic conversion, but the substrate, L-aspartate, is produced from petroleum, and fumarate production, which is also dependent on petroleum, is costly, resulting in low yields when replaced with other alternatives. Therefore, microbial fermentation offers an attractive alternative, providing a low-cost, abundant carbon source. Microorganisms naturally produce L-alanine using aminotransferases or alanine dehydrogenases. However, L-alanine production using naturally occurring strains is limited by low productivity.

[0005] Under these circumstances, the present inventors have made extensive efforts to develop an L-alanine-producing microorganism with a higher yield than conventional strains, and have found that a mutant strain obtained using Corynebacterium produces L-alanine in a high yield, thereby completing the present invention. Summary of the Invention [Problem to be solved by the invention]

[0006] One example of the present invention provides a mutant strain of the genus Corynebacterium, which is obtained by mutating the parent strain Corynebacterium glutamicum ATCC13869, and has improved L-alanine production ability compared to the parent strain.

[0007] Another example of the present invention is 1) Irradiating the parent strain Corynebacterium glutamicum ATCC 13869 to cause mutation; and 2) A method for producing a mutant strain having improved L-alanine production ability compared to the parent strain, comprising the steps of culturing the mutated strain of step 1) in a medium and selecting a mutant strain having improved L-alanine production ability.

[0008] Another embodiment of the present invention provides a composition for producing L-alanine, comprising the microorganism having improved L-alanine-producing ability.

[0009] Another embodiment of the present invention provides a method for producing L-alanine, comprising culturing the microorganism having improved L-alanine-producing ability in a medium.

[0010] Another embodiment of the present invention provides a use of the microorganism having improved L-alanine-producing ability for producing L-alanine. [Means for solving the problem]

[0011] This will be explained in detail below. Meanwhile, each description and embodiment disclosed in this application is also applicable to each other description and embodiment. In other words, all combinations of the various elements disclosed in this application fall within the scope of this application. Furthermore, the following specific description should not be construed as limiting the scope of this application. Furthermore, those skilled in the art will recognize or ascertain, using no more than routine experimentation, numerous equivalents to the specific aspects of this application described herein. Furthermore, such equivalents are intended to be encompassed by this application.

[0012] The present invention provides a microorganism with improved L-alanine-producing ability.

[0013] In the present invention, we conducted screening to find mutant strains that produce high levels of L-alanine. As a result, we confirmed that five mutant strains produced by irradiating Corynebacterium glutamicum ATCC13869 strain with radiation had improved L-alanine productivity compared to ATCC13869 strain.

[0014] The present invention provides a mutant strain of the genus Corynebacterium, which is obtained by mutating the parent strain Corynebacterium glutamicum ATCC13869, and has improved L-alanine production ability compared to the parent strain.

[0015] The microorganism may be a mutant strain of Corynebacterium glutamicum.

[0016] The Corynebacterium glutamicum mutant strains were Corynebacterium glutamicum CJ0230 strain (accession number KCCM13305P), Corynebacterium glutamicum CJ2011 strain (accession number KCCM13306P), Corynebacterium glutamicum CJ2015 strain (accession number KCCM13307P), Corynebacterium glutamicum CJ2057 strain (accession number KCCM13308P), and Corynebacterium glutamicum CJ2057 strain (accession number KCCM13309P). The bacterial strain may be any one selected from the group consisting of Lactobacillus casei strain CJ2064 (accession number KCCM13309P).

[0017] As used herein, the term "L-alanine" refers to an L-amino acid that is one of the essential amino acids and has the chemical formula "C3H7NO2."

[0018] As used herein, the term "productivity" means the ability to produce L-alanine and may be used interchangeably with "productivity." This can be confirmed by culturing an L-alanine-producing strain and then measuring the concentration of L-alanine in the culture medium.

[0019] The mutation may be carried out by various means known in the art, including physical or chemical mutagenesis methods. For example, a physical mutation method suitable for the present invention may be, but is not limited to, gamma ray or ultraviolet irradiation. Chemical mutation methods may include, but are not limited to, N-methyl-N'-nitro-N-nitrosoguanidine (NTG), diepoxybutane, ethyl methanesulfonate, mustard compounds, hydrazine, and nitrous acid.

[0020] The mutation for producing the mutant strain may be carried out by irradiation.

[0021] The radiation for inducing the mutation may be gamma rays or ultraviolet rays, preferably gamma rays.

[0022] The radiation may be administered at a dose of 3.0 kGy to 5.0 kGy, 3.1 kGy to 4.9 kGy, 3.2 kGy to 4.8 kGy, 3.3 kGy to 4.7 kGy, 3.4 kGy to 4.6 kGy, 3.5 kGy to 4.5 kGy, 3.6 kGy to 4.4 kGy, 3.7 kGy to 4.3 kGy, 3.8 kGy to 4.2 kGy, 3.9 kGy to 4.1 kGy, preferably 4.0 kGy per hour, but is not limited thereto.

[0023] The mutant strain may produce 1.5 times or more, 1.6 times or more, 1.7 times or more, or 1.5 to 3 times more L-alanine than the parent strain.

[0024] The mutant strain may have an L-alanine production ability that is 1.5 times or more improved, 1.6 times or more improved, 1.7 times or more improved, or even 1.5 to 3 times improved, compared to the parent strain.

[0025] The mutant strain may have an L-alanine productivity that is increased by 50% or more, 60% or more, 70% or more, 75% or more, 50% to 100%, 60% to 100%, 70% to 100%, 50% to 80%, 60% to 80%, 70% to 80%, or 75% compared to the parent strain.

[0026] The parent strain may be, but is not limited to, Corynebacterium glutamicum ATCC13869.

[0027] The present invention also provides 1) Irradiating the parent strain Corynebacterium glutamicum ATCC 13869 to cause mutation; and 2) A method for producing a mutant strain of the genus Corynebacterium having improved L-alanine production ability compared to the parent strain, comprising the steps of culturing the mutated strain of step 1) in a medium and selecting a mutant strain having improved L-alanine production ability.

[0028] The radiation may be gamma rays or ultraviolet rays, preferably gamma rays.

[0029] The radiation may be administered at a dose of 3.0 kGy to 5.0 kGy, 3.1 kGy to 4.9 kGy, 3.2 kGy to 4.8 kGy, 3.3 kGy to 4.7 kGy, 3.4 kGy to 4.6 kGy, 3.5 kGy to 4.5 kGy, 3.6 kGy to 4.4 kGy, 3.7 kGy to 4.3 kGy, 3.8 kGy to 4.2 kGy, 3.9 kGy to 4.1 kGy, preferably 4.0 kGy per hour, but is not limited thereto.

[0030] During mutagenesis, the parent strain is affected by a mutagen under conditions that leave a specific size of surviving individual bacteria. These conditions vary depending on the type of mutagen and depend on the amount of mutations that the mutagen induces in the surviving individual bacteria at a certain mortality rate (kill rate). For example, in the case of gamma rays, the mortality rate can be roughly 0.00001% to 20% of the starting population; in the case of NTG, the mortality rate can be roughly 10% to 50% of the starting population; and in the case of mutations induced by nitrite, the mortality rate can be 0.01% to 0.1% of the starting population, but is not limited to these.

[0031] The present invention also provides a composition for producing L-alanine, comprising a mutant strain of the genus Corynebacterium obtained by mutating the parent strain Corynebacterium glutamicum ATCC13869, and having improved L-alanine production ability compared to the parent strain.

[0032] The microorganism may be a mutant strain of Corynebacterium glutamicum.

[0033] The Corynebacterium glutamicum mutant strains were Corynebacterium glutamicum CJ0230 strain (accession number KCCM13305P), Corynebacterium glutamicum CJ2011 strain (accession number KCCM13306P), Corynebacterium glutamicum CJ2015 strain (accession number KCCM13307P), Corynebacterium glutamicum CJ2057 strain (accession number KCCM13308P), and Corynebacterium glutamicum CJ2057 strain (accession number KCCM13309P). The bacterial strain may be any one selected from the group consisting of Lactobacillus casei strain CJ2064 (accession number KCCM13309P).

[0034] In one embodiment of the present invention, mutant strains obtained by mutating the parent strain Corynebacterium glutamicum ATCC13869, including Corynebacterium glutamicum CJ0230 strain (accession number KCCM13305P), Corynebacterium glutamicum CJ2011 strain (accession number KCCM13306P), Corynebacterium glutamicum CJ2015 strain (accession number KCCM13307P), Corynebacterium glutamicum CJ2057 strain (accession number KCCM13308P), and Corynebacterium glutamicum It has been confirmed that the CJ2064 strain of L. glutamicum (accession number KCCM13309P) produces high concentrations of L-alanine. Therefore, a composition containing the mutant strain can be used to produce high concentrations of L-alanine.

[0035] The present invention also provides a method for producing L-alanine, which includes culturing in a medium a mutant strain of the genus Corynebacterium, which is obtained by mutating the parent strain Corynebacterium glutamicum ATCC13869 and has improved L-alanine-producing ability compared to the parent strain.

[0036] The method may additionally comprise, after the culturing step, recovering L-alanine from the cultured microorganism, the culture, or both.

[0037] As used herein, the term "culturing" refers to growing a microorganism under suitable artificially controlled environmental conditions. The method for culturing the microorganism of the present invention can be carried out using a culture method for Corynebacterium glutamicum that is widely known in the art. Specifically, examples of the culture method include, but are not limited to, batch culture, continuous culture, and fed-batch culture. Such various methods are disclosed, for example, in "Biochemical Engineering" (James M. Lee, Prentice-Hall International Editions, pp. 138-176, 1991).

[0038] As used herein, the term "culture" refers to a substance containing a medium in which microorganisms are growing or have completed growing under appropriately artificially controlled environmental conditions. In a narrow sense, the term "culture" does not include grown microorganisms, but in a broad sense it does. The "culture" includes medium components prepared for microbial culture as well as various substances released into the medium by the microorganisms during growth, specifically the target substance alanine.

[0039] The culture medium used for cultivation should be appropriately tailored to meet the requirements of the specific strain. Culture media for Corynebacterium strains are well known. For example, the microorganisms of the present application can be cultured in a conventional medium containing appropriate carbon sources, nitrogen sources, amino acids, vitamins, etc., under aerobic conditions, while controlling temperature and pH. Carbon sources include carbohydrates such as glucose, fructose, and sucrose, and amino acids such as glutamic acid and cysteine. Natural organic nutrient sources such as starch hydrolysates and molasses can be used, with carbohydrates such as glucose, fructose, and sterilized pretreated molasses (i.e., molasses converted to reducing sugars) being preferred. Various other carbon sources can be used in appropriate amounts without limitation, but are not limited to these. Nitrogen sources include inorganic nitrogen sources such as ammonia; amino acids such as glutamic acid and cysteine; and organic nitrogen sources such as peptone, meat extract, and yeast extract. These nitrogen sources can be used alone or in combination, but are not limited to these. The medium may contain, but is not limited to, phosphoric acid, potassium dihydrogen phosphate, dipotassium hydrogen phosphate, or the corresponding sodium-containing salts as a phosphorus source. Inorganic compounds may include magnesium sulfate, iron sulfate, manganese sulfate, and calcium chloride. Other ingredients may include amino acids, vitamins, and appropriate precursors. These media or precursors may be added to the culture in a batch or continuous manner, but are not limited to these.

[0040] During cultivation, compounds such as potassium hydroxide, ammonia, and phosphoric acid can be added to the culture in an appropriate manner to adjust the pH of the culture. Furthermore, during cultivation, antifoaming agents such as fatty acid polyglycol esters can be used to suppress foam formation. Oxygen or oxygen-containing gas can be injected into the culture to maintain an aerobic state. The culture temperature is 27°C to 37°C, specifically 30°C to 33°C. The cultivation period can be continued until the desired amount of useful substances is produced, specifically 20 to 120 hours.

[0041] The L-alanine recovery step may involve collecting the desired L-alanine from the culture medium, culture solution, or microorganism using a suitable method known in the art depending on the culture method. For example, the recovery step may be performed by one or more methods selected from, but not limited to, centrifugation, filtration, anion exchange chromatography, crystallization, HPLC, etc. The method for producing L-alanine may additionally include a purification step before, during, or after the recovery step. [Effects of the Invention]

[0042] The Corynebacterium mutant strain of the present invention is a microorganism whose L-alanine-producing ability is improved compared to that of the parent strain through the induction of mutation. Therefore, the method for producing L-alanine using the Corynebacterium mutant strain of the present invention can produce L-alanine with high efficiency and high yield. DETAILED DESCRIPTION OF THE INVENTION

[0043] The present invention will be described in more detail below with reference to the following examples, but these examples are for illustrative purposes only and are not intended to limit the scope of the present invention.

[0044] Example 1. Selection of mutant strains by artificial mutation method 1-1. Establishment of gamma-ray irradiation conditions to obtain mutant strains with improved L-alanine production To obtain a microbial mutant strain with improved L-alanine productivity, the following method was used to induce mutations in the microorganism.

[0045] To induce mutations, we applied gamma ray irradiation, a physical method. High-energy gamma rays emitted from the Co-60 radiation source flow along the chromosome, randomly inducing changes in base sequences such as base substitutions, deletions, and insertions. Because the level of base sequence changes is proportional to the intensity of the gamma ray irradiation, high-intensity gamma rays induce a high rate of mutations while also increasing the mortality rate (kill rate) of the strain. In order to ensure a sufficient level of high-quality mutant library with a high diversity of high mutation rates, we first established the gamma ray irradiation conditions for the corynebacterium strain to be irradiated.

[0046] Using a high-level gamma ray irradiation device from Nordion, located at the Advanced Radiation Research Institute of the Korea Atomic Energy Research Institute, 30 mL of seed medium culture solution with an absorbance 562 nm value of 6.09 was irradiated with gamma rays at intensities of 0, 0.5, 1, 2, 3, 4, 5, 7.5, and 10 kGy / hr for 1 hour, and then the activation medium was irradiated with 10 0 , 10 -1 , 10 -2 , 10 -3 , 10 -4 100 μl of each gamma-irradiated solution diluted at a dilution ratio of 0.5, 1, 2, 3, 4, 5, 7.5, and 10 kGy / hr was smeared on the plate, and the plate on which the gamma-irradiated solution was smeared was cultured in a static incubator at 30°C for 48 hours. The number of individual colonies formed was counted, and the mortality rate (killing rate) for each irradiation condition was calculated. The CFU / mL for gamma-irradiation conditions of 0, 0.5, 1, 2, 3, 4, 5, 7.5, and 10 kGy / hr was 3.6 x 10 8 , 6.9×10 7 , 9.8×10 6 , 2.7 × 10 5 , 2.1×10 3 , 3.0×10 1, 0, 0, 0, and the calculated mortality rates were 0%, 81.00467%, 97.31540%, 99.92589%, 99.99942%, 99.99999%, 100%, 100%, and 100%, respectively. Considering that gamma ray irradiation at an intensity of 5 kGy / hr resulted in complete death and failure to secure a colony, gamma ray irradiation at an intensity of 4 kGy / hr was established as the gamma ray irradiation condition for securing a high-quality mutant library with high mutation diversity through gamma ray irradiation of a corynebacterial strain.

[0047] 1-2. Selection of mutant strains with improved L-alanine production ability To identify a microorganism with high L-alanine productivity, we constructed a gamma-irradiation-based mutation library for Corynebacterium glutamicum ATCC13869 and screened for mutants with high L-alanine productivity. For gamma irradiation of ATCC13869, 50 mL of seed medium was cultured in a 500 mL flask in a shaking incubator at 30°C for 24 hours to obtain a culture with an Absorbance 562 nm value of 7.84. The culture was then diluted with the seed medium to an Absorbance 562 nm value of 6.10. Using a high-level gamma-irradiation system at the Advanced Radiation Research Institute of the Korea Atomic Energy Research Institute, 30 mL of the diluted solution was irradiated with 4 kGy / hr of gamma rays for 1 hour, and 100 μl of the irradiated stock solution was then spread onto 100 90 x 15 mm Petri dishes containing activated medium. The Petri dishes were then cultured in a static incubator at 30°C for 48 hours to obtain a total of approximately 40,000 individual colonies. To ensure the stability of the mutants, all individual colonies generated in all Petri dishes were collected and suspended in a 20% glycerol solution. The resulting suspension was then divided into 1 mL aliquots in 1.5 mL microtubes and stored in a cryogenic freezer at -80°C.

[0048] When screening for L-alanine high-producing mutants, a 20% glycerol suspension was taken out of the deep freezer and thawed at room temperature. -1 , 10 -2 , 10 -3 , 10-4 , 10 -5 After serial dilutions using 100 μl of each dilution, 100 μl of each dilution was smeared onto activation medium and cultured in a static incubator at 30°C for 24 hours to reactivate individual colonies. The reactivated individual colonies were inoculated into 96 deep-well plates containing 350 μl of production medium per well, equivalent to a 17% filling rate, using a Molecular Devices Qpix420 colony picker. The same plates were inoculated with ATCC13869 strains that had not been gamma-irradiated, in four wells per plate, to serve as a control for screening L-alanine high-producing mutants.

[0049] The 96-deep-well plates inoculated with the control strain and mutants were sealed using an Azenta Gas Permeable Seal Mark 2 and then cultured for 48 hours at 30°C and 1,000 rpm in an Infors-HT Multitron shaking incubator. After 48 hours of culture, the 96-deep-well plates were centrifuged for 20 minutes at 15°C and 4,000 rpm in an Eppendorf Centrifuge 5810R. Then, 100 μl of the culture supernatant from which the bacteria were isolated was transferred to a Corning 96-well Black Polystyrene Microplate using a Beckman Coulter Biomek i5 liquid handler for NIR spectrometry analysis. Next, the analysis was applied to NIR spectrometry, which was developed by the Analysis & Quality Department of CJ CheilJedang's Bio Technology Research Institute, to obtain individual analysis spectra for each well. This was then applied to a regression analysis prediction model with a coefficient of determination of 0.96 for the L-alanine concentration range of 0-20g / L, established based on culture samples whose L-alanine concentration had been quantified in advance by HPLC analysis, and selection logic for a 15% or greater increase in L-alanine concentration compared to the control group was applied to initially select 64 of the 10,304 mutant strains. The 64 selected strains were cultured in the same manner as above, and the top five strains with the highest L-alanine concentrations were finally selected.

[0050] The mutant strains obtained by the above method were named Corynebacterium glutamicum CJ0230, CJ2011, CJ2015, CJ2057, and CJ2064, and were deposited with the Korea Microorganism Collection, an international depository under the Budapest Treaty, on December 19, 2022, and assigned accession numbers KCCM13305P, KCCM13306P, KCCM13307P, KCCM13308P, and KCCM13309P, respectively.

[0051] The compositions of the media used in Examples 1 and 2 are as follows: <Activation medium> Beef extract 5g / L, Polypeptone 10g / L, Yeast extract 5g / L, Urea 2g / L, Sodium chloride (NaCl) 2.5g / L, Agar 20g / L, Glucose 10g / L, 10N sodium hydroxide (NaOH) 100μl / L

[0052] <Seed medium> Glucose (anhydrous glucose) 20g / L, polypeptone 10g / L, yeast extract 10g / L, ammonium sulfate [(NH4)2SO4] 10g / L, urea 1.5g / L, potassium phosphate monobasic (KH2PO4) 5.2g / L, potassium phosphate dibasic (K2HPO4) 10.7g / L, d-biotin 1.8mg / L, thiamine-HCl 9mg / L, CAPA 9mg / L, NCA 60mg / L, magnesium sulfate (MgSO4) 0.5g / L

[0053] <Production medium> Calcium carbonate (CaCO3) 30g / L, sucrose 57g / L, BM 6g / L, magnesium sulfate (MgSO4) 0.5g / L, (NH4)2SO4 50g / L, KH2PO4 1g / L, yeast extract 2g / L, ammonium acetate 6.28g / L, d-biotin 0.05mg / L, thiamine-HCl 0.1mg / L, MnSO4 6.7mg / L, FeSO4 10mg / L

[0054] Example 2. Evaluation of L-alanine productivity of L-alanine-producing mutant strains To confirm the L-alanine productivity of the Corynebacterium glutamicum strains CJ0230, CJ2011, CJ2015, CJ2057, and CJ2064 obtained in Example 1, they were cultured as follows.

[0055] Specifically, the parent strain, Corynebacterium glutamicum ATCC 13869, and the five mutants were each inoculated into a 250 ml corner baffle flask containing 25 ml of seed medium and cultured at 30°C for 20 hours with shaking at 200 rpm to obtain a seed culture. Then, 1 ml of the seed culture was inoculated into a 250 ml corner baffle flask containing 24 ml of the production medium and cultured at 30°C for 72 hours with shaking at 200 rpm to produce L-alanine. After the culture was completed, the L-alanine concentration in the culture medium was measured using high-performance liquid chromatography (HPLC) to determine the L-alanine production amount of each strain. The results are shown in Table 1 below.

[0056] [Table 1]

[0057] As a result, as shown in Table 1, the parent strain, Corynebacterium glutamicum ATCC13869, produced L-alanine at a concentration of 6.1 g / L. The mutant strains of Corynebacterium glutamicum according to the present invention, CJ0230, CJ2011, CJ2015, CJ2057, and CJ2064, produced L-alanine at concentrations of 10.7 g / L, 10.4 g / L, 9.9 g / L, 10.1 g / L, and 10.5 g / L, respectively, demonstrating that they had at least 162% higher L-alanine productivity than the parent strain.

[0058] The above results indicate that the five Corynebacterium glutamicum mutant strains according to the present invention can produce L-alanine with high efficiency and high yield. [Accession number]

[0059] Depository institution: Korea Center for Microorganisms (KCCM) Accession number: KCCM13305P Date of acceptance: 20221219

[0060] Depository institution: Korea Center for Microorganisms (KCCM) Accession number: KCCM13306P Date of acceptance: 20221219

[0061] Depository institution: Korea Center for Microorganisms (KCCM) Accession number: KCCM13307P Date of acceptance: 20221219

[0062] Depository institution: Korea Center for Microorganisms (KCCM) Accession number: KCCM13308P Date of acceptance: 20221219

[0063] Depository institution: Korea Center for Microorganisms (KCCM) Accession number: KCCM13309P Date of acceptance: 20221219

Claims

1. A mutant strain of the genus Corynebacterium obtained by mutating the parent strain Corynebacterium glutamicum ATCC13869, which has improved L-alanine productivity compared to the parent strain.

2. The mutant strain of the genus Corynebacterium according to claim 1, wherein the mutant strain of the genus Corynebacterium is Corynebacterium glutamicum.

3. The mutant strains of the genus Corynebacterium include Corynebacterium glutamicum CJ0230 strain (accession number KCCM13305P), Corynebacterium glutamicum CJ2011 strain (accession number KCCM13306P), Corynebacterium glutamicum CJ2015 strain (accession number KCCM13307P), Corynebacterium glutamicum CJ2016 strain (accession number KCCM13308P), Corynebacterium glutamicum CJ2015 strain (accession number KCCM13309P), Corynebacterium glutamicum CJ2014 strain (accession number KCCM13301P), Corynebacterium glutamicum CJ2015 strain (accession number KCCM13302P), Corynebacterium glutamicum CJ2016 strain (accession number KCCM13303P), Corynebacterium glutamicum CJ2015 strain (accession number KCCM13304P), Corynebacterium glutamicum CJ2016 strain (accession number KCCM13305P), Corynebacterium glutamicum CJ2016 strain (accession number KCCM13306P), Corynebacterium glutamicum CJ2016 strain (accession number KCCM13307P), Corynebacterium glutamicum CJ2016 strain (accession number KCCM13308P), Corynebacterium glutamicum CJ2016 strain (accession number KCCM13309P), Corynebacterium glutamicum CJ2016 strain (accession number KCCM13301P), Corynebacterium glutamicum CJ2016 strain (accession number KCCM13301P), Corynebacterium glutamicum CJ2016 2. The Corynebacterium mutant strain according to claim 1, which is any one selected from the group consisting of Corynebacterium glutamicum CJ2057 strain (accession number KCCM13308P) and Corynebacterium glutamicum CJ2064 strain (accession number KCCM13309P).

4. The mutant strain of Corynebacterium sp. according to claim 1 , wherein the mutation is induced by irradiation.

5. The Corynebacterium mutant strain according to claim 4 , wherein the radiation is gamma rays.

6. 5. The mutant strain of Corynebacterium according to claim 4, wherein the mutation is induced by irradiation at a dose of 3.5 kGy to 4.5 kGy per hour.

7. 2. The Corynebacterium mutant according to claim 1, wherein the mutant has an L-alanine productivity improved by 1.5 times or more compared to the parent strain, Corynebacterium glutamicum ATCC13869.

8. 1) irradiating the parent strain Corynebacterium glutamicum ATCC 13869 to cause mutation; and 2) A method for producing a mutant strain of Corynebacterium sp. having improved L-alanine production ability compared to the parent strain, comprising culturing the mutated strain of step 1) in a medium and selecting a mutant strain having improved L-alanine production ability.

9. The method for producing a mutant strain of the genus Corynebacterium according to claim 8 , wherein the radiation is gamma rays.

10. 9. The method for producing a mutant strain of the genus Corynebacterium according to claim 8, wherein the radiation is applied at a dose of 3.5 kGy to 4.5 kGy per hour.

11. The method for producing a mutant strain of the genus Corynebacterium according to claim 8, wherein the mutant strain of the genus Corynebacterium is Corynebacterium glutamicum.

12. The mutant strains of the genus Corynebacterium include Corynebacterium glutamicum CJ0230 strain (accession number KCCM13305P), Corynebacterium glutamicum CJ2011 strain (accession number KCCM13306P), Corynebacterium glutamicum CJ2015 strain (accession number KCCM13307P), Corynebacterium glutamicum CJ2016 strain (accession number KCCM13308P), Corynebacterium glutamicum CJ2015 strain (accession number KCCM13309P), Corynebacterium glutamicum CJ2014 strain (accession number KCCM13301P), Corynebacterium glutamicum CJ2015 strain (accession number KCCM13302P), Corynebacterium glutamicum CJ2016 strain (accession number KCCM13303P), Corynebacterium glutamicum CJ2015 strain (accession number KCCM13304P), Corynebacterium glutamicum CJ2016 strain (accession number KCCM13305P), Corynebacterium glutamicum CJ2016 strain (accession number KCCM13306P), Corynebacterium glutamicum CJ2016 strain (accession number KCCM13307P), Corynebacterium glutamicum CJ2016 strain (accession number KCCM13308P), Corynebacterium glutamicum CJ2016 strain (accession number KCCM13309P), Corynebacterium glutamicum CJ2016 strain (accession number KCCM13301P), Corynebacterium glutamicum CJ2016 strain (accession number KCCM13301P), Corynebacterium glutamicum CJ2016 9. The method for producing the mutant strain of the genus Corynebacterium according to claim 8, wherein the mutant strain is any one selected from the group consisting of Corynebacterium glutamicum CJ2057 strain (accession number KCCM13308P) and Corynebacterium glutamicum CJ2064 strain (accession number KCCM13309P).

13. 9. The method for producing a mutant strain of the genus Corynebacterium according to claim 8, wherein the mutant strain has an L-alanine productivity improved by 1.5 times or more compared to the parent strain Corynebacterium glutamicum ATCC13869.

14. A composition for producing L-alanine, comprising the Corynebacterium mutant strain according to claim 1.

15. The mutant strains of the genus Corynebacterium include Corynebacterium glutamicum CJ0230 strain (accession number KCCM13305P), Corynebacterium glutamicum CJ2011 strain (accession number KCCM13306P), Corynebacterium glutamicum CJ2015 strain (accession number KCCM13307P), Corynebacterium glutamicum CJ2016 strain (accession number KCCM13308P), Corynebacterium glutamicum CJ2015 strain (accession number KCCM13309P), Corynebacterium glutamicum CJ2014 strain (accession number KCCM13301P), Corynebacterium glutamicum CJ2015 strain (accession number KCCM13302P), Corynebacterium glutamicum CJ2016 strain (accession number KCCM13303P), Corynebacterium glutamicum CJ2015 strain (accession number KCCM13304P), Corynebacterium glutamicum CJ2016 strain (accession number KCCM13305P), Corynebacterium glutamicum CJ2016 strain (accession number KCCM13306P), Corynebacterium glutamicum CJ2016 strain (accession number KCCM13307P), Corynebacterium glutamicum CJ2016 strain (accession number KCCM13308P), Corynebacterium glutamicum CJ2016 strain (accession number KCCM13309P), Corynebacterium glutamicum CJ2016 strain (accession number KCCM13301P), Corynebacterium glutamicum CJ2016 strain (accession number KCCM13301P), Corynebacterium glutamicum CJ2016 15. The composition for producing L-alanine according to claim 14, wherein the bacterial strain is any one selected from the group consisting of Corynebacterium glutamicum CJ2057 strain (accession number KCCM13308P) and Corynebacterium glutamicum CJ2064 strain (accession number KCCM13309P).

16. A method for producing L-alanine, comprising culturing the Corynebacterium mutant strain of claim 1 in a medium.

17. The mutant strains of the genus Corynebacterium include Corynebacterium glutamicum CJ0230 strain (accession number KCCM13305P), Corynebacterium glutamicum CJ2011 strain (accession number KCCM13306P), Corynebacterium glutamicum CJ2015 strain (accession number KCCM13307P), Corynebacterium glutamicum CJ2016 strain (accession number KCCM13308P), Corynebacterium glutamicum CJ2015 strain (accession number KCCM13309P), Corynebacterium glutamicum CJ2014 strain (accession number KCCM13301P), Corynebacterium glutamicum CJ2015 strain (accession number KCCM13302P), Corynebacterium glutamicum CJ2016 strain (accession number KCCM13303P), Corynebacterium glutamicum CJ2015 strain (accession number KCCM13304P), Corynebacterium glutamicum CJ2016 strain (accession number KCCM13305P), Corynebacterium glutamicum CJ2016 strain (accession number KCCM13306P), Corynebacterium glutamicum CJ2016 strain (accession number KCCM13307P), Corynebacterium glutamicum CJ2016 strain (accession number KCCM13308P), Corynebacterium glutamicum CJ2016 strain (accession number KCCM13309P), Corynebacterium glutamicum CJ2016 strain (accession number KCCM13301P), Corynebacterium glutamicum CJ2016 strain (accession number KCCM13301P), Corynebacterium glutamicum CJ2016 17. The method for producing L-alanine according to claim 16, wherein the strain is any one selected from the group consisting of Corynebacterium glutamicum CJ2057 strain (accession number KCCM13308P) and Corynebacterium glutamicum CJ2064 strain (accession number KCCM13309P).

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