Crystal structure of the ferredoxin reductase component of carbazole 1,9a-dioxygenase from Janthinobacterium sp. J3.

Carbazole 1,9a-dioxygenase (CARDO), which consists of an oxygenase component and the electron-transport components ferredoxin (CARDO-F) and ferredoxin reductase (CARDO-R), is a Rieske nonheme iron oxygenase (RO). ROs are classified into five subclasses (IA, IB, IIA, IIB and III) based on their number of constituents and the nature of their redox centres. In this study, two types of crystal structure (type I and type II) were resolved of the class III CARDO-R from Janthinobacterium sp. J3 (CARDO-RJ3). Superimposition of the type I and type II structures revealed the absence of flavin adenine dinucleotide (FAD) in the type II structure along with significant conformational changes to the FAD-binding domain and the C-terminus, including movements to fill the space in which FAD had been located. Docking simulation of NADH into the FAD-bound form of CARDO-RJ3 suggested that shifts of the residues at the C-terminus caused the nicotinamide moiety to approach the N5 atom of FAD, which might facilitate electron transfer between the redox centres. Differences in domain arrangement were found compared with RO reductases from the ferredoxin-NADP reductase family, suggesting that these differences correspond to differences in the structures of their redox partners ferredoxin and terminal oxygenase. The results of docking simulations with the redox partner class III CARDO-F from Pseudomonas resinovorans CA10 suggested that complex formation suitable for efficient electron transfer is stabilized by electrostatic attraction and complementary shapes of the interacting regions.

Structural and digestion properties of potato starch modified using an efficient starch branching enzyme AqGBE.

Modified potato starch with slower digestion may aid the development of new starch derivatives with improved nutritional values, and strategies to increase nutritional fractions such as resistant starch (RS) are desired. In this study, a correspondence between starch structure and enzymatic resistance was provided based on the efficient branching enzyme AqGBE, and modified starches with different amylose content (Control, 100%; PS1, 90%; PS2, 72%; PS3, 32%; PS4, 18%) were prepared. Through SEM observation, NMR and X-ray diffraction analyses, we identified that an increased proportion of α-1,6-linked branches in potato starch changes its state of granule into large pieces with crystallinity. Molecular weight and chain-length distribution analysis showed a decrease of molecular weight (from 1.1 × 106 to 1.1 × 105 g/mol) without an obvious change of chain-length distribution in PS1, while PS2-4 exhibited an increased proportion of DP 6-12 with a stable molecular weight distribution, indicating a distinct model of structural modification by AqGBE. The enhancement of peak viscosity was related to increased hydrophobic interactions and pieces state of PS1, while the contents of SDS and RS in PS1 increased by 37.7 and 49.4%, respectively. Our result provides an alternative way to increase the RS content of potato starch by branching modification.

Heterologous expression of the gene for chlorite dismutase from Ideonella dechloratans is induced by an FNR-type transcription factor.

Regulation of the expression of the gene for chlorite dismutase (cld), located on the chlorate reduction composite transposon of the chlorate reducer Ideonella dechloratans, was studied. A 200 bp upstream sequence of the cld gene, and mutated and truncated versions thereof, was used in a reporter system in Escherichia coli. It was found that a sequence within this upstream region, which is nearly identical to the canonical FNR-binding sequence of E. coli, is necessary for anaerobic induction of the reporter gene. Anaerobic induction was regained in an FNR-deficient strain of E. coli when supplemented either with the fnr gene from E. coli or with a candidate fnr gene cloned from I. dechloratans. In vivo transcription of the suggested fnr gene of I. dechloratans was demonstrated by qRT-PCR. Based on these results, the cld promoter of I. dechloratans is suggested to be a class II-activated promoter regulated by an FNR-type protein of I. dechloratans. No fnr-type genes have been found on the chlorate reduction composite transposon of I. dechloratans, making anaerobic upregulation of the cld gene after a gene transfer event dependent on the presence of an fnr-type gene in the recipient.