ZnJ6 Is a Thylakoid Membrane DnaJ-Like Chaperone with Oxidizing Activity in Chlamydomonas reinhardtii.

Assembly of photosynthetic complexes is sensitive to changing light intensities, drought and pathogens, each of which induces a redox imbalance that requires the assistance of specific chaperones to maintain protein structure. Here we report a thylakoid membrane-associated DnaJ-like protein, ZnJ6 (Cre06.g251716.t1.2), in Chlamydomonas reinhardtii. The protein has four CXXCX(G)X(G) motifs that form two zinc fingers (ZFs). Site-directed mutagenesis (Cys > Ser) eliminates the ability to bind zinc. An intact ZF is required for ZnJ6 stability at elevated temperatures. Chaperone assays with recombinant ZnJ6 indicate that it has holding and oxidative activities. ZnJ6 is unable to reduce the disulfide bonds of insulin but prevents its aggregation in a reducing environment. It also assists in the reactivation of reduced denatured RNaseA, possibly by its oxidizing activity. ZnJ6 pull-down assays revealed interactions with oxidoreductases, photosynthetic proteins and proteases. In vivo experiments with a C. reinhardtii insertional mutant (∆ZnJ6) indicate enhanced tolerance to oxidative stress but increased sensitivity to heat and reducing conditions. Moreover, ∆ZnJ6 has reduced photosynthetic efficiency shown by the Chlorophyll fluorescence transient. Taken together, we identify a role for this thylakoid-associated DnaJ-like oxidizing chaperone that assists in the prevention of protein misfolding and aggregation, thus contributing to stress endurance, redox maintenance and photosynthetic balance.

Characterization of saltern based Streptomyces sp. and statistical media optimization for its improved antibacterial activity.

A moderately halotolerant Streptomyces strain, designated JAJ13 was characterized and a culture medium was statistically optimized to improve its antibacterial activity. Based on the phenotypic and molecular characteristics, strain JAJ13 was identified as a moderately halotolerant Streptomyces sp. JAJ13. Novelty of the strain JAJ13 in production of antibacterial compound was assessed by sequence analysis of KSα gene and LC-MS analysis of the active compound. Optimization of the culture medium for antibacterial compound production by the strain JAJ13 was performed with statistical methodology based on experimental designs. Initially, a starch based basal production medium was selected out of eight different production media screened for antibacterial compound production by Streptomyces sp. JAJ13. Plackett-Burman design was employed to screen the influential media components affecting the antibacterial compound production. Subsequently, statistical optimization of selected medium components was performed by employing the response surface methodology (RSM) with Box-Behnken design. The optimum initial level of CuSO4.5H2O, (NH4)2SO4 and K2HPO4 for the highest antibacterial activity was determined to be at 4.45 mg, 1.96 g, and 1.15 g in 1 L of distilled H2O, respectively. PBD and RSM guided design of experiments resulted in a maximum antibacterial activity of 23.37 ± 2.08 mm, which is a 78.8% increase in comparison with that obtained in the unoptimized medium. This study points the success of statistical model in developing an optimized production media for enhanced antibacterial compound production by Streptomyces sp. JAJ13.