1887

Abstract

In the current model of conjugal plasmid transfer in mycelium-forming streptomycetes, plasmid transfer by the FtsK-like TraB protein is followed by the subsequent spreading of the newly transferred plasmid within the neighbouring mycelial compartments. Several plasmid-encoded Spd proteins are involved in the plasmid spreading by an unknown mechanism. of the conjugative pSVH1 plasmid of was heterologously expressed in and , with a C-terminal His-tag-encoding sequence. Induction of -His expression affected viability in both species. The integral membrane protein SpdB2-His was eluted from the membrane fraction of with Triton X-100, and purified as a soluble protein by Ni-NTA affinity chromatography. Cross-linking experiments with glutaraldehyde showed that SpdB2-His formed oligomers. SpdB2-His had a nonspecific DNA-binding activity: while all types of dsDNA were bound, single-stranded M13-DNA was not recognized. The genes of the operon of pSVH1 were simultaneously expressed in with different affinity tags. While expression of StrepII-SpdB3 was not detected, Spd79-flag and SpdB2-His were localized in the membrane fraction of . In the absence of SpdB2, most of the Spd79-flag protein was found in the cytoplasmic fraction, indicating that SpdB2 affects localization of Spd79. Pulldown assays with StrepII-TraB protein of pSVH1 demonstrated that TraB interacted with SpdB2, suggesting that the septal DNA translocator TraB is also involved in intramycelial plasmid spreading.

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2007-09-01
2021-10-16
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