No abstract

Efficient lactic acid production from cane sugar molasses by Lactobacillus delbrueckii mutant Uc-3 in batch fermentation process is demonstrated. Lactic acid fermentation using molasses was not significantly affected by yeast extract concentrations. The final lactic acid concentration increased with increases of molasses sugar concentrations up to 190 g/liter. The maximum lactic acid concentration of 166 g/liter was obtained at a molasses sugar concentration of 190 g/liter with a productivity of 4.15 g/liter/h. Such a high concentration of lactic acid with high productivity from molasses has not been reported previously, and hence mutant Uc-3 could be a potential candidate for economical production of lactic acid from molasses at a commercial scale.

Heme oxygenase-2 (HO2) and -1 (HO1) catalyze heme degradation to biliverdin, CO, and iron, forming an essential link in the heme metabolism network. Tight regulation of the cellular levels and catalytic activities of HO1 and HO2 is important for maintaining heme homeostasis. HO1 expression is transcriptionally regulated; however, HO2 expression is constitutive. How the cellular levels and activity of HO2 are regulated remains unclear. Here, we elucidate the mechanism of post-translational regulation of cellular HO2 levels by heme. We find that, under heme-deficient conditions, HO2 is destabilized and targeted for degradation, suggesting heme plays a direct role in HO2 regulation. HO2 has three heme binding sites: one at its catalytic site, and the others at its two heme regulatory motifs (HRMs). We report that, in contrast to other HRM-containing proteins, the cellular protein level and degradation rate of HO2 are independent of heme binding to the HRMs. Rather, under heme deficiency, loss of heme binding to the catalytic site destabilizes HO2. Consistently, a HO2 catalytic site variant that is unable to bind heme exhibits a constant low protein level and an enhanced protein degradation rate compared to the wild-type HO2. Finally, HO2 is degraded by the lysosome through chaperone-mediated autophagy, distinct from other HRM-containing proteins and HO1, which are degraded by the proteasome. These results reveal a novel aspect of HO2 regulation and deepen our understanding of HO2’s role in maintaining heme homeostasis, paving the way for future investigation into HO2’s pathophysiological role in heme deficiency response.

The radical SAM enzyme, viperin, exerts a wide range of antiviral effects through both the synthesis of the antiviral nucleotide 3′-deoxy-3′,4′-didehydro-CTP (ddhCTP) and through its interactions with various cellular and viral proteins. Here we investigate the interaction of viperin with hepatitis C virus nonstructural protein 5A (NS5A) and the host sterol regulatory protein, vesicle-associated membrane protein A (VAP-33). NS5A and VAP-33 form part of the viral replication complex that is essential for replicating the RNA genome of the hepatitis C virus. Using transfected enzymes in HEK293T cells, we show that viperin binds independently to both NS5A and the C-terminal domain of VAP-33 (VAP-33C) and that this interaction is dependent on the proteins being colocalized to the ER membrane. Coexpression of VAP-33C and NS5A resulted in changes to the catalytic activity of viperin that depended upon viperin being colocalized to the ER membrane. The viperin-NS5A-VAP-33C complex exhibited the lowest specific activity, indicating that NS5A may inhibit viperin’s ability to synthesize ddhCTP. Coexpression of viperin with NS5A was also found to significantly reduce cellular NS5A levels, most likely by increasing the rate of proteasomal degradation. An inactive mutant of viperin, unable to bind the iron–sulfur cluster, was similarly effective at reducing cellular NS5A levels.

The Pseudomonas putida phenol-responsive regulator DmpR is a bacterial enhancer binding protein (bEBP) from the AAA + ATPase family. Even though it was discovered more than two decades ago and has been widely used for aromatic hydrocarbon sensing, the activation mechanism of DmpR has remained elusive. Here, we show that phenol-bound DmpR forms a tetramer composed of two head-to-head dimers in a head-to-tail arrangement. The DmpRphenol complex exhibits altered conformations within the C-termini of the sensory domains and shows an asymmetric orientation and angle in its coiled-coil linkers. The structural changes within the phenol binding sites and the downstream ATPase domains suggest that the effector binding signal is propagated through the coiled-coil helixes. The tetrameric DmpR-phenol complex interacts with the σ 54 subunit of RNA polymerase in presence of an ATP analogue, indicating that DmpR-like bEBPs tetramers utilize a mechanistic mode distinct from that of hexameric AAA + ATPases to activate σ 54-dependent transcription.