The Roles of the Various Cellulose Biosynthesis Operons in ATCC 23769.

Cellulose is the most abundant biopolymer on earth and offers versatile applicability in biotechnology. Bacterial cellulose, especially, is an attractive material because it represents pure microcrystalline cellulose. The cellulose synthase complex of acetic acid bacteria serves as a model for general studies on (bacterial) cellulose synthesis.

Expression of membrane-bound dehydrogenases from a mother of vinegar metagenome in Gluconobacter oxydans.

Acetic acid bacteria are well-known for their membrane-bound dehydrogenases rapidly oxidizing a variety of substrates in the periplasm. Since many acetic acid bacteria have not been successfully cultured in the laboratory yet, studying membrane-bound dehydrogenases directly from a metagenome of vinegar microbiota seems to be a promising way to identify novel variants of these enzymes. To this end, DNA from a mother of vinegar was isolated, sequenced, and screened for membrane-bound dehydrogenases using an in silico approach.

Characterization of membrane-bound dehydrogenases of Gluconobacter oxydans 621H using a new system for their functional expression.

Acetic acid bacteria are used in biotechnology due to their ability to incompletely oxidize a great variety of carbohydrates, alcohols, and related compounds in a regio- and stereo-selective manner. These reactions are catalyzed by membrane-bound dehydrogenases (mDHs), often with a broad substrate spectrum. In this study, the promoters of six mDHs of Gluconobacter oxydans 621H were characterized.

Identification of novel esterase-active enzymes from hot environments by use of the host bacterium Thermus thermophilus.

Functional metagenomic screening strategies, which are independent of known sequence information, can lead to the identification of truly novel genes and enzymes. Since E. coli has been used exhaustively for this purpose as a host, it is important to establish alternative expression hosts and to use them for functional metagenomic screening for new enzymes.

Importance of codB for new codA-based markerless gene deletion in Gluconobacter strains.

For the detailed molecular analysis, genomic modification, and application of acetic acid bacteria such as Gluconobacter in biotechnological processes, a simple markerless deletion system is essential. The available methods have either low efficiencies or their applicability is restricted to strains containing an upp mutation. We now developed a method based on counterselection by cytosine deaminase, encoded by the codA gene from Escherichia coli, in the presence of the fluorinated pyrimidine analogue 5-fluorocytosine (FC).

Characterization of membrane-bound dehydrogenases from Gluconobacter oxydans 621H via whole-cell activity assays using multideletion strains.

Gluconobacter oxydans, like all acetic acid bacteria, has several membrane-bound dehydrogenases, which oxidize a multitude of alcohols and polyols in a stereo- and regio-selective manner. Many membrane-bound dehydrogenases have been purified from various acetic acid bacteria, but in most cases without reporting associated sequence information. We constructed clean deletions of all membrane-bound dehydrogenases in G.

Deletion of pyruvate decarboxylase by a new method for efficient markerless gene deletions in Gluconobacter oxydans.

Gluconobacter oxydans, a biotechnologically relevant species which incompletely oxidizes a large variety of carbohydrates, alcohols, and related compounds, contains a gene for pyruvate decarboxylase (PDC). This enzyme is found only in very few species of bacteria where it is normally involved in anaerobic ethanol formation via acetaldehyde. In order to clarify the role of PDC in the strictly oxidative metabolism of acetic acid bacteria, we developed a markerless in-frame deletion system for strain G.

Metagenomic cellulases highly tolerant towards the presence of ionic liquids--linking thermostability and halotolerance.

Cellulose is an important renewable resource for the production of bioethanol and other valuable compounds. Several ionic liquids (ILs) have been described to dissolve water-insoluble cellulose and/or wood. Therefore, ILs would provide a suitable reaction medium for the enzymatic hydrolysis of cellulose if cellulases were active and stable in the presence of high IL concentrations.

A two-host fosmid system for functional screening of (meta)genomic libraries from extreme thermophiles.

A new cloning system is described, which allows the construction of large-insert fosmid libraries in Escherichia coli and the transfer of the recombinant libraries to the extreme thermophile Thermus thermophilus via natural transformation. Libraries are established in the thermophilic host by site-specific chromosomal insertion of the recombinant fosmids via single crossover or double crossover recombination at the T. thermophilus pyr locus.