MICROORGANISM BELONGING TO GENUS STAPHYLOCOCCUS PRODUCING ALLULOSE AND METHOD FOR PREPARING ALLULOSE USING THE SAME

Abstract

The present application relates to a microorganism that produces allulose and a method for preparing allulose using the same.

Claims

1. A composition for allulose production comprising a microorganism belonging to the genus Staphylococcus or a culture of the microorganism belonging to the genus Staphylococcus.

2. The composition according to claim 1, wherein the microorganism belonging to the genus Staphylococcus is any one selected from the group consisting of Staphylococcus agnetis, Staphylococcus argensis, Staphylococcus argenteus, Staphylococcus adettae, Staphylococcus aureus, Staphylococcus auricularis, Staphylococcus capitis, Staphylococcus caprae, Staphylococcus camosus, Staphylococcus chromogenes, Staphylococcus cohnii, Staphylococcus condiment, Staphylococcus comubiensis, Staphylococcus delphini, Staphylococcus devriesei, Staphylococcus edaphicus, Staphylococcus epidermidis, Staphylococcus equorum, Staphylococcus faecalis, Staphylococcus felis, Staphylococcus fleurettii, Staphylococcus gallinarum, Staphylococcus haemolyticus, Staphylococcus hominis, Staphylococcus hyicus, Staphylococcus intermedius, Staphylococcus kloosii, Staphylococcus leei, Staphylococcus lentus, Staphylococcus lugdunensis, Staphylococcus lutrae, Staphylococcus lyticans, Staphylococcus massiliensis, Staphylococcus microti, Staphylococcus muscae, Staphylococcus nepalensis, Staphylococcus pasteuri, Staphylococcus petrasii, Staphylococcus pettenkoferi, Staphylococcus piscifermentans, Staphylococcus pseudintermedius, Staphylococcus pseudolugdunensis, Staphylococcus rostri, Staphylococcus saccharolyticus, Staphylococcus saprophyticus, Staphylococcus schleifer, Staphylococcus schweitzeri, Staphylococcus sciuri, Staphylococcus simiae, Staphylococcus simulans, Staphylococcus stepanovicii, Staphylococcus succinus, Staphylococcus vitulinus, Staphylococcus warneri, and Staphylococcus xylosus.

3. The composition according to claim 1, wherein the microorganism belonging to the genus Staphylococcus is any one selected from the group consisting of Staphylococcus carnosus, Staphylococcus xylosus, Staphylococcus vitulinus, Staphylococcus epidermidis, Staphylococcus warneri, Staphylococcus haemolyticus, Staphylococcus intermedius, Staphylococcus saprophyticus, Staphylococcus cohnii, Staphylococcus muscae, Staphylococcus lentus, Staphylococcus chromogenes, Staphylococcus caprae, Staphylococcus auricularis, Staphylococcus gallinarum, Staphylococcus arlettae, Staphylococcus equorum, Staphylococcus kloosii, Staphylococcus delphini, and Staphylococcus pasteuri.

4. The composition according to claim 1, wherein the microorganism belonging to the genus Staphylococcus is nonpathogenic.

5. A method for preparing allulose, the method comprising bringing a microorganism belonging to the genus Staphylococcus or a culture of the microorganism belonging to the genus Staphylococcus into contact with fructose to convert the fructose to allulose.

6. The preparation method according to claim 5, wherein the microorganism belonging to the genus Staphylococcus is any one selected from the group consisting of Staphylococcus agnetis, Staphylococcus argensis, Staphylococcus argenteus, Staphylococcus arlettae, Staphylococcus aureus, Staphylococcus auricularis, Staphylococcus capitis, Staphylococcus caprae, Staphylococcus camosus, Staphylococcus chromogenes, Staphylococcus cohnii, Staphylococcus condiment, Staphylococcus cornubiensis, Staphylococcus delphini, Staphylococcus devriesei, Staphylococcus edaphicus, Staphylococcus epidermidis, Staphylococcus equorum, Staphylococcus faecalis, Staphylococcus felis, Staphylococcus fleurettii, Staphylococcus gallinarum, Staphylococcus haemolyticus, Staphylococcus hominis, Staphylococcus hyicus, Staphylococcus intermedius, Staphylococcus kloosii, Staphylococcus leei, Staphylococcus lentus, Staphylococcus lugdunensis, Staphylococcus lutrae, Staphylococcus lyticans, Staphylococcus massiliensis, Staphylococcus microti, Staphylococcus muscae, Staphylococcus nepalensis, Staphylococcus pasteuri, Staphylococcus petrasii, Staphylococcus pettenkoferi, Staphylococcus piscifermentans, Staphylococcus pseudintermedius, Staphylococcus pseudolugdunensis, Staphylococcus rostri, Staphylococcus saccharolyticus, Staphylococcus saprophyticus, Staphylococcus schleiferi, Staphylococcus schweitzeri, Staphylococcus sciuri, Staphylococcus simiae, Staphylococcus simulans, Staphylococcus stepanovicii, Staphylococcus succinus, Staphylococcus vitulinus, Staphylococcus warneri, and Staphylococcus xylosus.

7. The preparation method according to claim 5, wherein the microorganism belonging to the genus Staphylococcus is any one selected from the group consisting of Staphylococcus carnosus, Staphylococcus xylosus, Staphylococcus vitulinus, Staphylococcus epidermidis, Staphylococcus warneri, Staphylococcus haemolyticus, Staphylococcus intermedius, Staphylococcus saprophyticus, Staphylococcus cohnii, Staphylococcus muscae, Staphylococcus lentus, Staphylococcus chromogenes, Staphylococcus caprae, Staphylococcus auricularis, Staphylococcus gallinarum, Staphylococcus arlettae, Staphylococcus equorum, Staphylococcus kloosii, Staphylococcus delphini, and Staphylococcus pasteuri.

8. The preparation method according to claim 5, which further comprises recovering converted allulose.

9. The preparation method according to claim 5, wherein the converting fructose to allulose is performed at a temperature of 40° C. to 70° C.

10. Use of a microorganism belonging to the genus Staphylococcus or a culture of the microorganism belonging to the genus Staphylococcus for allulose production.

Description

BRIEF DESCRIPTION OF DRAWINGS

[0059] FIG. 1 is a diagram for confirming the production of allulose by Staphylococcus delphini KCTC3592:

[0060] FIG. 2 is a diagram illustrating the phylogenetic tree of nonpathogenic Staphylococcus strains; and

[0061] FIG. 3 is a diagram illustrating the phylogenetic tree of 20 nonpathogenic Staphylococcus strains.

DETAILED DESCRIPTION OF THE INVENTION

[0062] The best mode for carrying out the invention will be described in detail as follows. Meanwhile, each description and embodiment disclosed in the present application may also be applied to other descriptions and embodiments. That is, all combinations of various elements disclosed in the present application fall within the scope of the present application. Further, the scope of the present application is not limited by the specific description below.

[0063] In order to achieve the above object, the present application provides a composition for allulose production comprising a microorganism belonging to the genus Staphylococcus or a culture of the microorganism belonging to the genus Staphylococcus.

EXAMPLES

[0064] Hereinafter, the present application will be described in more detail with reference to Examples. However, these Examples are for illustrative purposes only, and the scope of the present application is not limited to these Examples.

Example 1: Confirmation of Allulose Production by Microorganism Belonging to Genus Staphylococcus

[0065] In order to confirm the ability of microorganisms belonging to the genus Staphylococcus to produce allulose, 32 nonpathogenic microorganisms belonging to the genus Staphylococcus were selected, and it was confirmed whether 20 microorganisms belonging to the genus Staphylococcus among these produced allulose from D-fructose.

[0066] Specifically, 2 species of Staphylococcus carnosus (KCTC3580, KACC13250), 2 species of Staphylococcus xylosus (KCTC3342, KACC16180), 4 species of Staphylococcus vitulinus (KACC15803, KACC15804, KACC15805, KACC13211), Staphylococcus delphini (KCTC3592), Staphylococcus equorum (KCTC3589), Staphylococcus epidermidis (KCTC1917), Staphylococcus cohnii (KCTC3574), Staphylococcus chromogenes (KCTC3579), Staphylococcus caprae (KCTC3583), Staphylococcus warneri (KCTC3340), Staphylococcus lentus (KCTC3577), Staphylococcus muscae (KCTC3576), Staphylococcus saprophyticus (KCTC3345), Staphylococcus pasteuri (KCTC13167), Staphylococcus intermedius (KCTC3344), Staphylococcus arlettae (KCTC3588), Staphylococcus kloosii (KCTC3590), Staphylococcus haemolyticus (KCTC3341), Staphylococcus gallinarum (KCTC3585), and Staphylococcus auricularis (KCTC3584) were distributed from the Korean Collection for Type Cultures (KTCT) and the Korean Agricultural Culture Collection (KACC).

[0067] Each of the microorganisms was inoculated into Tryptic Soy Broth (peptone 17 g/L, soytone 3 g/L, psicose 10 g/L, NaCl 5 g/L, K.sub.2HPO.sub.4 2.5 g/L, agar 15 g/L) to which allulose was added at 1%, and cultured at 30° C. or 37° C. for 18 hours. Thereafter, the cultured microorganisms were recovered, washed with 0.85% (w/v) NaCl, and then used to perform the whole-cell conversion reaction.

[0068] To the microorganism at a concentration of 20% (w/w), 50 mM potassium phosphate buffer (pH 7.5) to which D-fructose was added at 1% (w/w) was added and suspended, and the suspension was subjected to the conversion reaction at 55° C. for 12 hours.

[0069] The supernatant of the conversion reaction product was subjected to HPLC analysis to confirm the allulose production. HPLC analysis was performed at a temperature of 80° C. and a flow velocity of 0.6 mL/min using water as a mobile phase solvent and a Refractive Index Detector (Agilent 1260 RID) of HPLC (Agilent, USA) equipped with Aminex HPX-87C column (BIO-RAD). The allulose conversion rate was calculated as the ratio of the weight of allulose produced after the reaction to the weight of substrate (D fructose) before the reaction (allulose concentration (12 h reaction)/initial fructose concentration (1 wt %)).

[0070] As a result, it has been confirmed that the 20 microorganisms belonging to the genus Staphylococcus all produce allulose from D-fructose (Table and FIG. 1).

TABLE-US-00001 TABLE 1 Name of strain Conversion rate (%) Staphylococcus delphini KCTC3592 29.8 Staphylococcus carnosus KCTC3580 26.2 Staphylococcus carnosus KACC13250 25.7 Staphylococcus equorum KCTC3589 24.5 Staphylococcus epidermidis KCTC1917 15.8 Staphylococcus cohnii KCTC3574 10.4 Staphylococcus xylosus KCTC3342  8.0 Staphylococcus xylosus KACC16180  8.0 Staphylococcus chromogenes KCTC3579  7.8 Staphylococcus caprae KCTC3583  5.3 Staphylococcus vitulinus KACC15803  3.3 Staphylococcus vitulinus KACC15804  3.4 Staphylococcus warneri KCTC3340  2.7 Staphylococcus lentus KCTC3577  2.9 Staphylococcus vitulinus KACC15805  2.6 Staphylococcus muscae KCTC3576  2.3 Staphylococcus pasteuri KCTC13167  1.8 Staphylococcus intermedius KCTC3344  1.7 Staphylococcus saprophyticus KCTC3345  1.6 Staphylococcus arlettae KCTC3588  1.2 Staphylococcus vitulinus KACC13211  0.9 Staphylococcus kloosii KCTC3590  0.8 Staphylococcus haemolyticus KCTC3341  0.5 Staphylococcus gallinarum KCTC3585  0.4 Staphylococcus auricularis KCTC3584  0.3

Example 2: Confirmation of 16S rRNA Similarity of Microorganisms Belonging to Genus Staphylococcus

Example 2-1: Confirmation of 16S rRNA Similarity of Nonpathogenic Microorganisms Belonging to Genus Staphylococcus

[0071] Microorganisms classified as nonpathogenic (biosafety level 1) microorganisms belonging to the genus Staphylococcus in the present Example are as follows: Staphylococcus argensis, Staphylococcus capitis, Staphylococcus devriesei, Staphylococcus faecalis, Staphylococcus sciuri, Staphylococcus hominis, Staphylococcus lugdunensis, Staphylococcus microti, Staphylococcus piscifermentans, Staphylococcus schweitzeri, Staphylococcus simulans, Staphylococcus succinus, Staphylococcus arlettae, Staphylococcus auricularis, Staphylococcus caprae, Staphylococcus camosus, Staphylococcus chromogenes, Staphylococcus cohnii, Staphylococcus delphini, Staphylococcus epidermidis, Staphylococcus equorum, Staphylococcus gallinarum, Staphylococcus haemolyticus, Staphylococcus intermedius, Staphylococcus kloosii, Staphylococcus lentus, Staphylococcus muscae, Staphylococcus pasteuri, Staphylococcus saprophyticus, Staphylococcus vitulinus, Staphylococcus warneri, and Staphylococcus xylosus.

[0072] The sequence homology between strains was analyzed using the Clustal omega program (https://www.ebi.ac.uk/Tools/msa/clustalo) based on the 16s rRNA sequences of 31 microorganisms among the nonpathogenic microorganisms belonging to the genus Staphylococcus except Staphylococcus faecalis, of which the 16s rRNA sequence was not identified. The representative 16s rRNA sequences of the respective strains were confirmed from the NCBI database.

[0073] As a result, it has been confirmed that the homology of 16s rRNA sequences of the microorganisms belonging to the genus Staphylococcus known as nonpathogenic microorganisms between the strains is 92.78% or more in all cases (FIG. 2). The homology of the 31 microorganisms belonging to the genus Staphylococcus was in most cases 95% or more, and the homology was 92.78% to 95.21% in the case of Staphylococcus argensis.

[0074] This is a result supporting that the microorganisms belonging to the genus Staphylococcus classified as nonpathogenic microorganisms all have high genetic connectivity.

Example 2-2: Confirmation of 16S rRNA Similarity of 20 Nonpathogenic Microorganisms Belonging to Genus Staphylococcus

[0075] The sequence homology was analyzed using the Clustal omega program (https://www.ebi.ac.uk/Tools/msa/clustalo) based on the 16s rRNA sequences of the 20 microorganisms belonging to the genus Staphylococcus of which the ability to produce allulose was confirmed in Example 1 above. The representative 16s rRNA sequences of the respective strains were confirmed via the NCBI database.

[0076] As a result, it has been confirmed that the homology of 16s rRNA sequences of the 20 microorganisms belonging to the genus Staphylococcus of which the ability to produce allulose has been confirmed is 95.97% or more.

[0077] From the Examples above, it has been confirmed that nonpathogenic microorganisms belonging to the genus Staphylococcus all have high genetic connectivity between species, and the microorganisms belonging to the genus Staphylococcus that have such high genetic connectivity exhibit the activity of producing allulose from fructose.

[0078] Based on the above description, it will be understood by those skilled in the art that the present disclosure may be implemented in a different specific form without changing the technical spirit or essential characteristics thereof.

[0079] Therefore, it should be understood that the above embodiment is not limitative, but illustrative in all aspects. The scope of the disclosure is defined by the appended claims rather than by the description preceding them, and therefore all changes and modifications that fall within metes and bounds of the claims or equivalents of such metes and bounds are therefore intended to be embraced by the claims.