Purification and Characterization of a New Thermostable Laccase from Bacillus licheniformis O12 Using One-Step Affinity Chromatography and Its Potential for Decolorization

Arzu Öztürk Kesebir; Melda Şişecioğlu; Ahmet Adıgüzel; Deryanur Kılıç; Ömer İrfan Küfrevioğlu

Makale detayları

Purification and Characterization of a New Thermostable Laccase from Bacillus licheniformis O12 Using One-Step Affinity Chromatography and Its Potential for Decolorization

Yazarlar:

Arzu Öztürk Kesebir | Melda Şişecioğlu | Ahmet Adıgüzel | Deryanur Kılıç | Ömer İrfan Küfrevioğlu

Yayıncı:

Igdir University

Ana Başlıklar:

Biyokimya ve Hücre Biyolojisi (Diğer) | Biyolojik Olarak Aktif Moleküller | Proteinler ve Peptitler

Dergi:

Iğdır Üniversitesi Fen Bilimleri Enstitüsü Dergisi Vol.14 Issue 1, pp.333-344

Belge Tipi:

Makale

Yayın Tarihi:

1/2024

Belge Dili:

İngilizce

Document ID (Org No.):

2001311749

pISSN:

2536-4618

Anahtar kelimeler:

Özet:

 Laccases are copper-containing enzymes that can oxidize a wide variety of substrates. Thanks to this feature of laccase, some dyes that cause environmental pollution can be decolorized. Some bacteria, such as Bacillus licheniformis, naturally produce the enzyme laccase. A new affinity column was tested in this study. For this purpose, the extracellular laccase sepharose 4B-L-tyrosine-ρ-aminobenzoic acid produced by bacteria grown in suitable media was isolated by affinity chromatography method. Its purity was checked by SDS-PAGE method. The decolorization effect of some dyestable in textile wastewater of laccase isolated from B. lichenisformis O12 by affinity column was investigated. No mediator was used in this procedure. .As a result, laccase was purified 4.82-fold purification with a yield of 38.3 respectively, The molecular weight of the purified enzyme was determined as 70 kDa by the SDS-PAGE method. The enzyme showed optimum activity at pH 4.0 and temperature 92C. The enzyme was found to retain 100 activity even after 12 hours of incubation at 60C and 92C. The kinetic parameters were determined with laccase substrates such as ABTS, 2,6-DMP, and guaiacol. The purified laccase was decolorized with varied efficiencies such as 35 of Reactive black, 31 of Acid black 1, 28 of Methylene blue, and 15 of Acid red 27 without the use of any redox mediators. These properties of B. licheniformis O12 laccase enzyme make it a potential candidate enzyme for use in various biotechnological and industrial applications.

 Laccases are copper-containing enzymes that can oxidize a wide variety of substrates. Thanks to this feature of laccase, some dyes that cause environmental pollution can be decolorized. Some bacteria, such as Bacillus licheniformis, naturally produce the enzyme laccase. A new affinity column was tested in this study. For this purpose, the extracellular laccase sepharose 4B-L-tyrosine-ρ-aminobenzoic acid produced by bacteria grown in suitable media was isolated by affinity chromatography method. Its purity was checked by SDS-PAGE method. The decolorization effect of some dyestable in textile wastewater of laccase isolated from B. lichenisformis O12 by affinity column was investigated. No mediator was used in this procedure. .As a result, laccase was purified 4.82-fold purification with a yield of 38.3 respectively, The molecular weight of the purified enzyme was determined as 70 kDa by the SDS-PAGE method. The enzyme showed optimum activity at pH 4.0 and temperature 92C. The enzyme was found to retain 100 activity even after 12 hours of incubation at 60C and 92C. The kinetic parameters were determined with laccase substrates such as ABTS, 2,6-DMP, and guaiacol. The purified laccase was decolorized with varied efficiencies such as 35 of Reactive black, 31 of Acid black 1, 28 of Methylene blue, and 15 of Acid red 27 without the use of any redox mediators. These properties of B. licheniformis O12 laccase enzyme make it a potential candidate enzyme for use in various biotechnological and industrial applications.

 Laccases are copper-containing enzymes that can oxidize a wide variety of substrates. Thanks to this feature of laccase, some dyes that cause environmental pollution can be decolorized. Some bacteria, such as Bacillus licheniformis, naturally produce the enzyme laccase. A new affinity column was tested in this study. For this purpose, the extracellular laccase sepharose 4B-L-tyrosine-ρ-aminobenzoic acid produced by bacteria grown in suitable media was isolated by affinity chromatography method. Its purity was checked by SDS-PAGE method. The decolorization effect of some dyestable in textile wastewater of laccase isolated from B. lichenisformis O12 by affinity column was investigated. No mediator was used in this procedure. .As a result, laccase was purified 4.82-fold purification with a yield of 38.3 respectively, The molecular weight of the purified enzyme was determined as 70 kDa by the SDS-PAGE method. The enzyme showed optimum activity at pH 4.0 and temperature 92C. The enzyme was found to retain 100 activity even after 12 hours of incubation at 60C and 92C. The kinetic parameters were determined with laccase substrates such as ABTS, 2,6-DMP, and guaiacol. The purified laccase was decolorized with varied efficiencies such as 35 of Reactive black, 31 of Acid black 1, 28 of Methylene blue, and 15 of Acid red 27 without the use of any redox mediators. These properties of B. licheniformis O12 laccase enzyme make it a potential candidate enzyme for use in various biotechnological and industrial applications.

 Lakkazlar çok çeşitli substratları oksitleyebilen bakır içeren enzimlerdir. Lakkazın bu özelliği sayesinde çevre kirliliğine neden olan bazı boyarmaddelerin renk giderimi yapılabilmektedir. Bacillus licheniformis gibi bazı bakteriler doğal olarak lakkaz enzimini üretir. Çalışmada lakkaz saflaştırması için yeni bir afinite kolonu test edilmiştir. Bu amaçla uygun ortamda yetiştirilen bakterilerin ürettiği hücre dışı lakkaz sefaroz 4B-L-tirozin-ρ-aminobenzoik asit afinite kromatografisi yöntemiyle izole edildi. Saflığı SDS-PAGE yöntemiyle kontrol edildi. B. lichenisformis O12den afinite kolonu ile izole edilen lakkazın tekstil atık suyundaki bazı boyarmaddelerin renk giderici etkisi araştırıldı. Bu prosedürde herhangi bir mediyatör kullanılmadı. Sonuç olarak lakkaz 38,3 verimle 4,82 kat saflaştırıldı. Saflaştırılan enzimin moleküler ağırlığı SDS-PAGE yöntemiyle 70 kDa olarak belirlendi. Enzim, pH 4.0da ve 92C sıcaklıkta optimum aktivite gösterdi. Enzimin, 60C ve 92Cde 12 saatlik inkübasyondan sonra bile 100 aktivitesini koruduğu görüldü. Kinetik parametreler ABTS, 2,6-DMP ve guaiacol gibi lakkaz substratları ile belirlendi. Ortama sadece izole edilen lakkaz eklenerek, herhangi bir redoks aracısı kullanılmadan 35 Reaktif siyah, 31 Asit siyahı 1, 28 Metilen mavisi ve 15 Asit kırmızısı 27 boyar maddelerinde renk giderimi elde edildi. B. licheniformis O12 lakkaz enziminin bu özellikleri, onu çeşitli biyoteknolojik ve endüstriyel uygulamalarda kullanım için potansiyel bir aday enzim haline getirmektedir.

Makale detayları

Purification and Characterization of a New Thermostable Laccase from Bacillus licheniformis O12 Using One-Step Affinity Chromatography and Its Potential for Decolorization

Yazarlar:

Arzu Öztürk Kesebir | Melda Şişecioğlu | Ahmet Adıgüzel | Deryanur Kılıç | Ömer İrfan Küfrevioğlu

Yayıncı:

Igdir University

Ana Başlıklar:

Biyokimya ve Hücre Biyolojisi (Diğer) | Biyolojik Olarak Aktif Moleküller | Proteinler ve Peptitler

Dergi:

Iğdır Üniversitesi Fen Bilimleri Enstitüsü Dergisi Vol.14 Issue 1, pp.333-344

Belge Tipi:

Makale

Yayın Tarihi:

1/2024

Belge Dili:

İngilizce

Document ID (Org No.):

2001311749

pISSN:

2536-4618

Bacillus licheniformis

Laccase

Affinity chromatography

Characterization

Decolorization

PurificationBacillus licheniformis

Lakkaz

Afinite Kromatografisi

Karakterizasyon

Renk giderimi

Saflaştırma

Sayı

Makale detayları

Yazarlar: Arzu Öztürk Kesebir | Melda Şişecioğlu | Ahmet Adıgüzel | Deryanur Kılıç | Ömer İrfan Küfrevioğlu

Yayıncı: Igdir University

Ana Başlıklar: Biyokimya ve Hücre Biyolojisi (Diğer) | Biyolojik Olarak Aktif Moleküller | Proteinler ve Peptitler

Dergi: Iğdır Üniversitesi Fen Bilimleri Enstitüsü Dergisi Vol.14 Issue 1, pp.333-344

Belge Tipi: Makale

Yayın Tarihi: 1/2024

Belge Dili: İngilizce

Document ID (Org No.): 2001311749

pISSN: 2536-4618

Sayı

Yazarlar:

Arzu Öztürk Kesebir | Melda Şişecioğlu | Ahmet Adıgüzel | Deryanur Kılıç | Ömer İrfan Küfrevioğlu

Yayıncı:

Igdir University

Ana Başlıklar:

Biyokimya ve Hücre Biyolojisi (Diğer) | Biyolojik Olarak Aktif Moleküller | Proteinler ve Peptitler

Dergi:

Iğdır Üniversitesi Fen Bilimleri Enstitüsü Dergisi Vol.14 Issue 1, pp.333-344

Belge Tipi:

Makale

Yayın Tarihi:

1/2024

Belge Dili:

İngilizce

Document ID (Org No.):

2001311749

pISSN:

2536-4618