The Genomes of the Non-Clearing-Zone-Forming and Natural-Rubber- Degrading Species Gordonia polyisoprenivorans and Gordonia westfalica Harbor Genes Expressing Lcp Activity in Streptomyces Strains

Abstract

The latex-clearing protein (Lcp K30 ) from the rubber-degrading bacterium Streptomyces sp. strain K30 is involved in the cleavage of poly(cis-1,4-isoprene), yielding isoprenoid aldehydes and ketones. Lcp homologues have so far been detected in all investigated clearing-zone-forming rubber-degrading bacteria. Internal degenerated oligonucleotides derived from lcp genes of Streptomyces sp. strain K30 (lcp K30 ), Streptomyces coelicolor strain A3(2), and Nocardia farcinica strains IFM10152 and E1 were applied in PCR to investigate whether lcp homologues occur also in the non-clearing-zone-forming rubber-utilizing bacteria Gordonia polyisoprenivorans strains VH2 and Y2K, Gordonia alkanivorans strain 44187, and Gordonia westfalica strain Kb1, which grow adhesively on rubber. The 1,230-and 1,224-bp lcp-homologous genes from G. polyisoprenivorans strain VH2 (lcp VH2 ) and G. westfalica strain Kb1 (lcp Kb1 ) were obtained after screening genomic libraries by degenerated PCR amplification, and their translational products exhibited 50 and 52% amino acid identity, respectively, to Lcp K30 . Recombinant lcp VH2 and lcp Kb1 harboring cells of the non-rubber-degrading Streptomyces lividans strain TK23 were able to form clearing zones and aldehydes on latex overlay-agar plates, thus indicating that lcp VH2 and lcp Kb1 encode functionally active proteins. Analysis by gel permeation chromatography demonstrated lower polymer concentrations and molecular weights of the remaining polyisoprenoid molecules after incubation with these recombinant S. lividans strains. Reverse transcription-PCR analysis demonstrated that lcp VH2 was transcribed in cells of G. polyisoprenivorans strain VH2 cultivated in the presence of poly(cis-1,4-isoprene) but not in the presence of sodium acetate. Anti-Lcp K30 immunoglobulin Gs, which were raised in this study, were rather specific for Lcp K30 and did not cross-react with Lcp VH2 and Lcp Kb1 . A lcp VH2 disruption mutant was still able to grow with poly(cis-1,4-isoprene) as sole carbon source; therefore, lcp VH2 seems not to be essential for rubber degradation in G. polyisoprenivorans.

Involvement of Two Latex-Clearing Proteins during Rubber Degradation and Insights into the Subsequent Degradation Pathway Revealed by the Genome Sequence of Gordonia polyisoprenivorans Strain VH2

“…The Lcp gene products comprise a Tat signal peptide at the N terminus. This postulate fits well with observations that were made for non-clear-zone-forming S. lividans strain TK23 heterologously expressing lcp1 VH2 (21), and one could assume that Lcp is secreted by strain VH2. Furthermore, secretion of Lcp via the Tat pathway was already demonstrated in the clear-zone-forming Streptomyces sp.…”

“…Furthermore, after incubation of liquid medium with cells of this recombinant strain, the remaining polyisoprenoid molecules showed lower concentrations and molecular weights. Using reverse transcription-PCR analysis, we demonstrated that this gene is transcribed in the presence of rubber but not in the presence of sodium acetate (21). However, after this gene was deleted, the resulting mutant (G. polyisoprenivorans strain M71) showed growth behavior only slightly different from that of the wild type, as it grew only a little slower but reached the same cell density (Fig.…”

“…This kind of cleavage was also postulated for other adhesively and non-adhesively growing bacteria (18,89). Since heterologous expression of lcp1 VH2 enables S. lividans strain TK23 to form clear zones and aldehydes on latex overlay-agar plates, the remaining polyisoprenoid molecules are of lower polymer concentration and molecular weight (21), and a deletion of both Lcpencoding genes of strain VH2 resulted in an IR-negative phenotype, we postulate that the lcp genes are responsible for this oxidative cleavage of rubber. The Lcp gene products comprise a Tat signal peptide at the N terminus.…”

“…Furthermore, lcp VH2 transcripts in VH2 cultures grown on poly(cis-1,4-isoprene) as the sole carbon and energy source were detected by reverse transcription-PCR analysis, whereas no transcripts were detected after growth in the presence of sodium acetate. Nevertheless, a lcp VH2 disruption did not result in a negative rubber-degrading phenotype (21).…”

Biodegradation of Natural Rubber Latex of Calotropis procera by Two Endophytic Fungal Species

“…Many other reports have been published on the biodegradation of natural rubber hydrocarbon as a sole carbon source by bacteria [15][16][17][18][19][20][21][22], fungi [13,23,24]. Several microbes were used to biodegrade Calotropis Procera latex [25,26].…”