Pyruvate Decarboxylase is Less Effective Than That of for Ethanol Production in Metabolically Engineered sp. PCC6803.

To produce bioethanol from model cyanobacteria such as , a two gene cassette consisting of genes encoding pyruvate decarboxylase (PDC) and alcohol dehydrogenase (ADH) are required to transform pyruvate first to acetaldehyde and then to ethanol. However the partition of pyruvate to ethanol comes at a cost, a reduction in biomass and pyruvate availability for other metabolic processes. Hence strategies to divert flux to ethanol as a biofuel in are of interest.

The quaternary structure of Thermus thermophilus aldehyde dehydrogenase is stabilized by an evolutionary distinct C-terminal arm extension.

Aldehyde dehydrogenases (ALDH) form a superfamily of dimeric or tetrameric enzymes that catalyze the oxidation of a broad range of aldehydes into their corresponding carboxylic acids with the concomitant reduction of the cofactor NAD(P) into NAD(P)H. Despite their varied polypeptide chain length and oligomerisation states, ALDHs possess a conserved architecture of three domains: the catalytic domain, NAD(P) binding domain, and the oligomerization domain. Here, we describe the structure and function of the ALDH from Thermus thermophilus (ALDH) which exhibits non-canonical features of both dimeric and tetrameric ALDH and a previously uncharacterized C-terminal arm extension forming novel interactions with the N-terminus in the quaternary structure.

Evaluation of the ethanol tolerance for wild and mutant strains by flow cytometry.

Flow cytometry was used to evaluate the effect of initial ethanol concentrations on cyanobacterial strains of PCC 6803 [wild-type (WT), and ethanol producing recombinants (UL 004 and UL 030)] in batch cultures. Ethanol recombinants, containing one or two metabolically engineered cassettes, were designed towards the development of an economically competitive process for the direct production of bioethanol from microalgae through an exclusive autotrophic route. It can be concluded that the recombinant UL 030 containing two copies of the genes per genome was the most tolerant to ethanol.

Analysis and comparative genomics of R997, the first SXT/R391 integrative and conjugative element (ICE) of the Indian Sub-Continent.

The aim of this study was to analyse R997, the first integrative and conjugative element (ICE) isolated from the Indian Sub-Continent, and to determine its relationship to the SXT/R391 family of ICEs. WGS of Escherichia coli isolate AB1157 (which contains R997) was performed using Illumina sequencing technology. R997 context was assessed by de novo assembly, gene prediction and annotation tools, and compared to other SXT/R391 ICEs.

SXT/R391 Integrative and Conjugative Elements (ICEs) Encode a Novel 'Trap-Door' Strategy for Mobile Element Escape.

Integrative conjugative elements (ICEs) are a class of bacterial mobile elements that have the ability to mediate their own integration, excision, and transfer from one host genome to another by a mechanism of site-specific recombination, self-circularisation, and conjugative transfer. Members of the SXT/R391 ICE family of enterobacterial mobile genetic elements display an unusual UV-inducible sensitization function which results in stress induced killing of bacterial cells harboring the ICE. This sensitization has been shown to be associated with a stress induced overexpression of a mobile element encoded conjugative transfer gene, orf43, a traV homolog.