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Summary: Natural transformation increases the genetic diversity of bacteria, but is costly and must be strictly controlled. We previously found that deletion of ccpA, a key regulator of carbon catabolite repression (CCR), reduced transformation efficiency of Streptococcus oligofermentans, the current work further investigated the regulatory mechanisms of CcpA. The competence operon comCDE is subjected to basal and autoregulatory transcription. A luciferase reporter detected a transcriptional readthrough (TRT) from the upstream tRNAArg into the comCDE operon, which was induced by L‐arginine. Insertion of the Escherichia coli T1T2 terminator downstream of tRNAArg abolished TRT, and reduced the basal comCDE transcription by 77% and also the transformation efficiency. Deletion of ccpA increased tRNAArg TRT and tRNAArg‐comCDE polycistronic transcript by twofold. An in vitro transcription assay determined that CcpA promoted the transcription termination of tRNAArg TRT, and RNA EMSA and SPR assays detected equal binding affinity of CcpA to both the RNA and DNA of tRNAArg. These results indicate that CcpA controls the basal comCDE transcription by post‐transcriptional actions. Overexpression of comDE or its phospho‐mimicking mutant comDED58E reduced transformation efficiency, indicating that excessive ComE impairs competence development. CCR‐regulated competence was further confirmed by higher tRNAArg TRT but lower transformation efficiency in galactose than in glucose. [ABSTRACT FROM AUTHOR]

A titanium dioxide supported VPO(VPO/TiO2) catalyst for NH3-SCR de-NOx was prepared. The NH3-SCR catalytic activity of VPO/TiO2 was tested and a corresponding mechanism was investigated by Density Functional Theory and in situ FTIR spectra. The results showed that the catalytic activity of VPO/TiO2 was the highest when the molar ratio of P to V was 1/5, weight percentage of active ingredient was 10 wt.% and calcination temperature was 400 °C. The de-NOx efficiency of 0.1VP(1/5)O/TiO2 calcined at 400 °C was above 98% at temperature range from 180 to 400 °C. The V2P2O15H12 cluster was constructed and the adsorption of NO and NH3 on the active site of VPO/TiO2 was investigated by density functional theory (DFT). The simulation results showed that NO could be chemisorbed on the O2 and O3 site of the V2P2O15H12 cluster, and the corresponding adsorption energy was − 74.95 kJ·mol−1 and − 47.30 kJ·mol−1 respectively. The adsorption energy of NH3 adsorption on O1, O2 and O3 site is − 95.88 kJ·mol−1, − 230.80 kJ·mol−1 and − 78.45 kJ·mol−1. Moreover, the electric charge transformation of H on O2 site is 0.589e, which is higher than that on O1 and O3 site. Accordingly, the NH3-SCR de-NOx reaction would occur more easily on the O2 site than on the O1 and O3 site. The simulated results and the in situ FTIR spectra showed that the reduction of NO by NH3 over VPO/TiO2 followed the E–R mechanism and L–H mechanism. [ABSTRACT FROM AUTHOR]

Strictosidine synthase (STR), the gate enzyme for monoterpenoid indole alkaloid biosynthesis, catalyzes the Pictet-Spengler reaction (PSR) of various tryptamine derivatives with secologanin assisted by "indole sandwich" stabilization. Continuous exploration with β-methyltryptamine (IPA) stereoselectively delivered the C6-methylstrictosidines and C6-methylvincosides by enzymatic and nonenzymatic PSR, respectively. Unexpectedly, the first "nonindole sandwich" binding mode was witnessed by the X-ray structures of STR1-ligand complexes. Site-directed mutagenesis revealed the critical cryptic role of the hydroxyl group of Tyr151 in IPA biotransformation. Further computational calculations demonstrated the adjustable IPA position in STR1 upon the binding of secologanin, and Tyr151-OH facilitates the productive PSR binding mode via an advantageous hydrogen-bond network. Further chemo-enzymatic manipulation of C6-methylvincosides successfully resulted in the discovered antimalarial framework (IC 50 = 0.92 μM).

A cell of gram-negative bacteria is surrounded by two layers of membrane, the inner membrane and the outer membrane. Proteins are the major composition of outer membrane. Many outer membrane proteins carry a trans-membrane β-barrel structure that formed by multiple anti-parallel β-strands connected with hydrogen bonds. These proteins can act as porins, transporters, enzymes, receptors, virulence factors and structural proteins. Therefore, their correct folding and membrane integration are important for the survival of gram-negative bacteria. Most β-barrel outer membrane proteins could be easily expressed recombinantly and refolded in vitro under certain conditions. The in vitro folding processes could be monitored and investigated through many ways, which makes outer membrane proteins become a model system to study the effects of abiotic and biological factors on the folding of membrane proteins. In this article, the research progress on the in vitro refolding of outer membrane proteins are reviewed from the aspects of refolding methods, the factors that affect folding processes and experimental methods. Finally, the research prospects in this field are discussed.