Abstract

Two types of enzyme for oxidative cleavage of poly(cis-1,4-isoprene) are known. One is rubber oxygenase (RoxA) that is secreted by Xanthomonas sp. 35Y and a few other Gram-negative rubber-degrading bacteria during growth on polyisoprene. RoxA was studied in the past and the recently solved structure showed a structural relationship to bacterial cytochrome c peroxidases (Seidel,J. et al., 2013, PNAS 110:13833-38). The other enzyme is latex-clearing-protein (Lcp) that is secreted by rubber degrading actinomycetes but Lcp has not yet been purified. Here, we expressed Lcp of Streptomyces sp. K30 in a ΔroxA background of Xanthomonas, sp. 35Y and purified native (untagged) Lcp. The specific activities of Lcp and RoxA were 0.70 U/mg and 0.48 U/mg, respectively. Lcp differed from RoxA in the absence of heme groups and other characteristics. Notably, Lcp degraded polyisoprene via endo-cleavage to tetra-(C20) and higher oligo-isoprenoids with aldehyde and keto end groups whereas RoxA employed an exo-cleavage type mechanism to give the main end product 12-oxo-4,8-dimethyltrideca-4,8-diene-1-al (ODTD). RoxA was able to cleave isolated Lcp-derived oligo-isoprenoid molecules to ODTD. Inhibitor studies, spectroscopic investigations and metal analysis gave no indication for the presence of iron, other metals or co-factors in Lcp. Our results suggest that Lcp could be a member of the growing group of co-factor-independent oxygenases and differs in the cleavage mechanism from heme-dependent RoxA. In conclusion, RoxA and Lcp represent two different answers to the same biochemical problem, the cleavage of polyisoprene, a polymer that has carbon-carbon double bonds as the only functional groups for enzymatic attack.