Publication: Functional significance of fructan biosynthesis in halophilic archaea and eubacteria
Abstract
Fruktanlar, fruktozdan oluşan homopolimerleri kapsayan bir polisakkarit grubunun genel adıdır ve mikrobiyal olanları fruktoz birimleri arasındaki bağlara göre levan (β-2,6 bağlı) ve inülinler (β-2,1 bağlı) olarak iki temel gruba ayrılırlar. Fruktanların doğada pek çok farklı bakteri, birçok çiçekli bitki ve kısmen bazı mantarlar tarafından üretildiği gösterilmiştir. Doğada aşırı tuzlu ortamlarda yaşayan halofilik bakterilerin de fruktan ürettiği ilk kez araştırma grubumuz tarafından gösterilmiş ve o zamandan bu yana Halomonas smyrnensis AAD6 bakterisi tarafından üretilen levan pek çok tıbbi, farmasötik, gıda ve kimya uygulamasında potansiyelini ortaya koymuştur. Bu bakterinin levan üretim mekanizmasının daha iyi anlaşılması için bu tez çalışması kapsamında ilk olarak mikroorganizmanın levan üretiminden sorumlu levansukraz enzimi (HsLsc) rekombinant olarak üretilip saflaştırılmış ve karakterize edilmiştir. Çalışmanın ikinci kısmında Çamaltı Tuzlası (İzmir, Türkiye) kaynaklı halotolerant Bacillus licheniformis OK12 suşunun ürettiği levan karakterize edilmiş ve genom analizi ile levanın potansiyeli metabolik yolaklara etkisi irdelenmiştir. Tez çalışmasının üçüncü kısmında ise aşırı tuzlu ortamlarda baskın olarak bulunan halofilik arkelerin de genomlarında fruktoziltransferaz genlerini taşıdığının fark edilmesi üzerine fruktan üretme kapasiteleri araştırılmıştır. Bunun için Tuz Gölü’nden (Türkiye) izole edilen ve kültür koleksiyonundan sipariş edilen Halomicrobium, Haloferax ve Natronococcus cinslerine üye toplam yedi farklı halofilik arke suşu ile sukroz varlığında polisakkaritler üretilmiş ve hepsinin baskın olarak inülin tipi fruktanlar ürettiği anlaşılmıştır. Bu çalışma, fruktanların bitkiler ve bakteriler dışında arkeler tarafından da üretildiğini gösteren ilk araştırma olma niteliği taşımaktadır.
Fructans are a group of polysaccharides majorly composed of fructose and those from microorganisms are classified into two main groups as levans (β-2,6 linked) and inulins (β-2,1 linked) according to the linkages between the fructose units. It has been shown that fructans are produced by a wide variety of bacteria, some flowering plants and fungi. It has been shown for the first time by our research group that fructans are also accumulated by halophilic bacteria which thrive in extremely saline habitats, and since then, levan from Halomonas smyrnensis AAD6, Halomonas levan, has shown its potential in many medical, pharmaceutical, food and chemical applications. To better understand the levan synthesis mechanism of this bacterium, the levansucrase enzyme (HsLsc) responsible for levan production by the microorganism has been recombinantly produced, purified and characterized. In the second part of the thesis, levan from halotolerant Bacillus licheniformis OK12 strain Çamaltı Saltern (İzmir, Turkey) was characterized and the putative contribution of its hydrolysis products to metabolic pathways was assessed. In the third part of this thesis, fructan synthesis capacities of halophilic Archaea were assessed since putative fructosyltransferase genes were found in their genomes. For this, seven different halophilic archaeal strains belonging to the genera of Halomicrobium, Haloferax and Natronococcus were either isolated from Tuz Lake (Turkey) or purchased from a culture collection were grown in the presence of sucrose and polysaccharides produced were determined to be mainly inulin-type fructans. This work represents the first research in the literature showing that apart from plants and Eubacteria, fructans are also synthesized by Archaea.
Fructans are a group of polysaccharides majorly composed of fructose and those from microorganisms are classified into two main groups as levans (β-2,6 linked) and inulins (β-2,1 linked) according to the linkages between the fructose units. It has been shown that fructans are produced by a wide variety of bacteria, some flowering plants and fungi. It has been shown for the first time by our research group that fructans are also accumulated by halophilic bacteria which thrive in extremely saline habitats, and since then, levan from Halomonas smyrnensis AAD6, Halomonas levan, has shown its potential in many medical, pharmaceutical, food and chemical applications. To better understand the levan synthesis mechanism of this bacterium, the levansucrase enzyme (HsLsc) responsible for levan production by the microorganism has been recombinantly produced, purified and characterized. In the second part of the thesis, levan from halotolerant Bacillus licheniformis OK12 strain Çamaltı Saltern (İzmir, Turkey) was characterized and the putative contribution of its hydrolysis products to metabolic pathways was assessed. In the third part of this thesis, fructan synthesis capacities of halophilic Archaea were assessed since putative fructosyltransferase genes were found in their genomes. For this, seven different halophilic archaeal strains belonging to the genera of Halomicrobium, Haloferax and Natronococcus were either isolated from Tuz Lake (Turkey) or purchased from a culture collection were grown in the presence of sucrose and polysaccharides produced were determined to be mainly inulin-type fructans. This work represents the first research in the literature showing that apart from plants and Eubacteria, fructans are also synthesized by Archaea.