Phenol is a common environmental contaminant. The purpose of this study was to isolate phenol-degrading microorganisms from wastewater in the sections of the Chinese Medicine Manufactory. The phenol-degrading Acinetobacter lwoffii NL1 was identified based on a combination of biochemical characteristics and 16S rRNA genes. To analyze the molecular mechanism, the whole genome of A. lwoffii NL1 was sequenced, yielding 3499 genes on one circular chromosome and three plasmids. Enzyme activity analysis showed that A. lwoffii NL1 degraded phenol via the ortho-cleavage rather than the meta-cleavage pathway. Key genes encoding phenol hydroxylase and catechol 1,2-dioxygenase were located on a megaplasmid (pNL1) and were found to be separated by mobile genetic elements; their function was validated by heterologous expression in Escherichia coli and quantitative real-time PCR. A. lwoffii NL1 could degrade 0.5 g/L phenol within 12 h and tolerate a maximum of 1.1 g/L phenol, and showed resistance against multiple antibiotics and heavy metal ions. Overall, this study shows that A. lwoffii NL1 can be potentially used for efficient phenol degradation in heavy metal wastewater treatment.

Bacterial type IV secretion system (T4SS) belongs to a growing class of evolutionarily conserved transporters that translocate DNA and proteins into a wide variety of organisms including bacterial and eukaryotic cells. Archetypal is the Agrobacterium tumefaciens VirB/D4 T4SS that transfers oncogenic T-DNA to various eukaryotic cells, which is transferred as a nucleoprotein T-complex with VirD2 as the pilot protein. As a derivative of plasmid conjugation systems, the VirB/D4 T4SS can also transfer certain mobilizable plasmids and bacterial proteins like VirE2 and VirF, although it is unknown how the membrane-bound T4SS recruits different transfer substrates. Here, we show that a cytoplasmic VirD2-binding protein (VBP) is involved in the recruitment of the T-complex to the energizing components of the T4SS, including VirD4, VirB4, and VirB11. VBP is also important for the recruitment of a conjugative plasmid to a different transfer system independent of VirB/D4. These data indicate that VBP functions as a previously unrecognized recruiting protein that helps couple nucleoprotein substrates to the appropriate transport sites for conjugative DNA transfers. VBP has three functionally redundant homologs, and similar homologs can be found in different bacterial genomes, suggesting a previously uncharacterized class of proteins involved in conjugative DNA transfers.

Agrobacterium tumefaciens is the agent that causes crown gall tumor disease on more than 140 species of dicotyledonous plants. Chemotaxis of A. tumefaciens toward the wound sites of the host plant is the first step to recognize the host. CheW is a coupling protein that bridges the histidine kinase CheA and the chemoreceptors to form the chemotaxis core signaling complex and plays a crucial role in the assembly and function of the large chemosensory array. Unlike all previously reported chemotaxis systems, A. tumefaciens has only one major che operon but two cheW homologs (atu2075 as cheW₁ and atu2617 as cheW₂) on unlinked loci. The in-frame deletion of either cheW gene significantly affects A. tumefaciens chemotaxis but does not abolish the chemotaxis, unless both cheW genes were deleted. The effect of cheW₂ deletion on the chemotaxis is more severe than that of cheW₁ deletion. Either CheW can interact with CheA and couple it to the cell poles. The promoter activity of cheW₂ is always higher than that of cheW₁ under all of the tested conditions. When two cheW genes were adjusted to the same expression level by using the identical promoter, the difference between the effects of two CheW proteins on the chemotaxis still existed. Therefore, we envision that both the different molecular ratio of two CheW proteins in cell and the different affinities of two CheW proteins with CheA and chemoreceptors result in the efficiency difference of two CheW proteins in functioning in the large chemosensory array.