RT Journal Article SR Electronic(1) A1 Oetermann, Sylvia A1 Jongsma, Rense A1 Coenen, Anna A1 Keller, Jeanne A1 Steinbüchel, AlexanderYR 2019 T1 LcpRVH2 – regulating the expression of latex-clearing proteins in Gordonia polyisoprenivorans VH2 JF Microbiology, VO 165 IS 3 SP 343 OP 354 DO https://doi.org/10.1099/mic.0.000755 PB Microbiology Society, SN 1465-2080, AB Gordonia polyisoprenivorans VH2 harbours two latex clearing proteins, which are responsible for the cleavage of poly(cis-1,4-isoprene) into oligoisoprenes, thereby allowing growth in presence of, e.g. natural rubber. A gene coding for a putative regulator of the TetR-family (lcpR VH2) is located 131 bp upstream of lcp1 VH2. We heterologously expressed lcpR VH2 in Escherichia coli , and purified and characterized the protein with respect to its ability to bind to the operator region of lcp1 VH2. LcpRVH2 forms a dimer in its native state. The size of the dimer was determined to be 52.7 kDa by size exclusion chromatography, whereas the calculated size of a monomer was 24.1 kDa. Electrophoretic mobility shift assays (EMSAs) with the purified protein revealed a shift upon binding to the intergenic region between lcpR VH2 and lcp1 VH2. Within this region, an inverted repeat was identified in silico, probably being the binding site of LcpRVH2. This binding sequence was confirmed by a DNase I footprinting assay. A shift also occurred in EMSAs with this 44 bp sequence only. Interestingly, no regulator was detected upstream of the second lcp (lcp2 VH2). Therefore, we performed EMSA studies with LcpRVH2 and the putative operator region upstream of lcp2 VH2, and discovered by DNase I footprinting another binding sequence upstream of lcp2 VH2. Hence, we concluded that LcpRVH2 binds the operator region of both lcps and, most likely, regulates their expression in G. polyisoprenivorans VH2., UL https://www.microbiologyresearch.org/content/journal/micro/10.1099/mic.0.000755