Mosaic Cooperativity in Slow Polypeptide Topological Isomerization Revealed by Residue-Specific NMR Thermodynamic Analysis.

Slow polypeptide conformational changes on time scales of >1 s are generally assumed to be highly cooperative two-state transitions, reflecting the high energy barrier. However, few experimental characterizations have tested the validity of this assumption. We performed residue-specific NMR thermodynamic analysis of the 27-residue lantibiotic peptide, nukacin ISK-1, to characterize the isomerization between two topological states on the second time scale. Unexpectedly, the thermal transition behaviors were distinct among peptide regions, indicating that the topological isomerization process is a mosaic of different degrees of cooperativity. The conformational change path between the two NMR structures was deduced by a targeted molecular dynamics simulation. The unique side-chain threading motions through the monosulfide rings are the structural basis of the high energy barrier, and the nonlocal interactions in the hydrophobic core are the structural basis of the cooperativity. Taken together, we provide an energetic description of the topological isomerization of nukacin ISK-1.

High Improvement in Lactic Acid Productivity by New Alkaliphilic Bacterium Using Repeated Batch Fermentation Integrated with Increased Substrate Concentration.

Optically pure lactic acid (LA) is an important chemical platform that has a wide range of industrial and biotechnological applications. Improved parameters for cost effective LA production are of great interest for industrial developments. In the present study, an alkaliphilic lactic acid bacterium, BoM 1-2, was selected among 369 newly obtained bacterial isolates. It was characterized using API 50 CHL kit and identified as Enterococcus hirae BoM 1-2 by 16S rRNA gene sequence analysis. Efficient polymer-grade L-lactic acid production was achieved at pH 9.0 and 40°C. In batch fermentation strategy using 20 g L-1 glucose, 19.6 g L-1 lactic acid was obtained with volumetric productivity of 2.18 g L-1 h-1. While using 100 g L-1 glucose, 96.0 g L-1 lactic acid was obtained with volumetric productivity of 1.07 g L-1 h-1. The highest lactic acid concentration of 180.6 g L-1 was achieved in multipulse fed batch strategy with volumetric productivity of 0.65 g L-1 h-1. To achieve higher productivity, repeated fermentation processes were applied using the two different strategies. In the first strategy, the lactic acid productivity was increased from 1.97 g L-1 h-1 to 4.48 g L-1 h-1 when the total of 10 repeated runs were carried out using 60 g L-1 glucose, but lactic acid productivity decreased to 2.95 g L-1 h-1 using 100 g L-1 glucose. In second strategy, repeated fermentation coupled with gradual increase in glucose concentration from 40 to 100 g L-1 was conducted for 24 runs. A dramatic increase in LA productivity up to 39.9 g L-1 h-1 (18-fold compared to first run) was achieved using 40 g L-1 glucose while volumetric productivity ranging between 24.8 and 29.9 g L-1 h-1 was achieved using 60-100 g L-1 glucose.

The lantibiotic nukacin ISK-1 exists in an equilibrium between active and inactive lipid-II binding states.

The lantibiotic nukacin ISK-1 exerts antimicrobial activity through binding to lipid II. Here, we perform NMR analyses of the structure of nukacin ISK-1 and the interaction with lipid II. Unexpectedly, nukacin ISK-1 exists in two structural states in aqueous solution, with an interconversion rate on a time scale of seconds. The two structures differ in the relative orientations of the two lanthionine rings, ring A and ring C. Chemical shift perturbation induced by the titration of lipid II reveals that only one state was capable of binding to lipid II. On the molecular surface of the active state, a multiple hydrogen-bonding site formed by amino acid residues in the ring A region is adjacent to a hydrophobic surface formed by residues in the ring C region, and we propose that these sites interact with the pyrophosphate moiety and the isoprene chain of the lipid II molecule, respectively.

Enhanced Bioethanol Production from Potato Peel Waste Via Consolidated Bioprocessing with Statistically Optimized Medium.

In this study, an extensive screening was undertaken to isolate some amylolytic microorganisms capable of producing bioethanol from starchy biomass through Consolidated Bioprocessing (CBP). A total of 28 amylolytic microorganisms were isolated, from which 5 isolates were selected based on high α-amylase and glucoamylase activities and identified as Candida wangnamkhiaoensis, Hyphopichia pseudoburtonii (2 isolates), Wickerhamia sp., and Streptomyces drozdowiczii based on 26S rDNA and 16S rDNA sequencing. Wickerhamia sp. showed the highest ethanol production (30.4 g/L) with fermentation yield of 0.3 g ethanol/g starch. Then, a low cost starchy waste, potato peel waste (PPW) was used as a carbon source to produce ethanol by Wickerhamia sp. Finally, in order to obtain maximum ethanol production from PPW, a fermentation medium was statistically designed. The effect of various medium ingredients was evaluated initially by Plackett-Burman design (PBD), where malt extracts, tryptone, and KH2PO4 showed significantly positive effect (p value < 0.05). Using Response Surface Modeling (RSM), 40 g/L (dry basis) PPW and 25 g/L malt extract were found optimum and yielded 21.7 g/L ethanol. This study strongly suggests Wickerhamia sp. as a promising candidate for bioethanol production from starchy biomass, in particular, PPW through CBP.

Bactericidal activity of nukacin ISK-1: an alternative mode of action.

We previously reported bacteriostatic action of nukacin ISK-1 against Bacillus subtilis JCM 1465(T). Here, we found its bactericidal activity against Micrococcus luteus DSM 1790 and Staphylococcus simulans 22, showing decrease in cell viability, cell lysis, and dissipation of the membrane potential. Moreover, leakage of small molecules such as K(+), suggested the formation of small-sized or specific K(+)-conducting-pores by nukacin ISK-1.