Secretion of Biologically Active Heterologous Oxalate Decarboxylase (OxdC) inLactobacillus plantarumWCFS1 Using Homologous Signal Peptides

Abstract

Current treatment options for patients with hyperoxaluria and calcium oxalate stone diseases are limited and do not always lead to sufficient reduction in urinary oxalate excretion. Oxalate degrading bacteria have been suggested for degrading intestinal oxalate for the prevention of calcium oxalate stone. Here, we reported a recombinant Lactobacillus plantarum WCFS1 (L. plantarum) secreting heterologous oxalate decarboxylase (OxdC) that may provide possible therapeutic approach by degrading intestinal oxalate. The results showed secretion and functional expression of OxdC protein in L. plantarum driven by signal peptides Lp_0373 and Lp_3050. Supernatant of the recombinant strain containing pLp_0373sOxdC and pLp_3050sOxdC showed OxdC activity of 0.05 U/mg and 0.02 U/mg protein, while the purified OxdC from the supernatant showed specific activity of 18.3 U/mg and 17.5 U/mg protein, respectively. The concentration of OxdC protein in the supernatant was 8–12 μg/mL. The recombinant strain showed up to 50% oxalate reduction in medium containing 10 mM oxalate. In conclusion, the recombinant L. plantarum harboring pLp_0373sOxdC and pLp_3050sOxdC can express and secrete functional OxdC and degrade oxalate up to 50% and 30%, respectively.

Observation of superoxide production during catalysis of Bacillus subtilis oxalate decarboxylase at pH 4

“…It accumulates in the cell wall of Bacillus subtilis and can be secreted by many basidiomycetous and ascomycetous fungi [11, 12]. OxDC has garned attention due to its potentially numerous applications ranging from remediation of oxalate scaling in the wood and paper industry [13–16], bioengineering of crop plants for fungal resistance and lower oxalate content [17, 18], diagnostics and sensing of oxalate [19–21], bioengineering of probiotic gut bacteria to release OxDC in the intestine [22], and as a dietary supplement for the degradation of excess oxalate in the stomach [23, 24]. Despite these efforts, significant questions remain about the details of the enzymatic mechanism of OxDC, and in particular about how modifications of the enzyme can direct its chemistry away from decarboxylase to oxidase activities [25].…”

Mobile group II intron based gene targeting in Lactobacillus plantarum WCFS1

“…Similarly a novel Ll.LtrB intron expression plasmid (pLpACD4C) created in this study can be useful for gene targeting in L. plantarum WCFS1. Previously, heterologous oxalate decarboxylase was expressed in L. plantarum and preliminary results suggested that usage of recombinant L. plantarum prevent the calcium oxalate stone formation in rats . The genetically modified lactic acid bacterium developed in this study with its biological containment status will be used as probiotic tools to deliver oxalate decarboxylase into the human intestine for the prevention of hyperoxaluria and renal stone disease in future.…”

“…Recently, recombinant Lactobacillus plantarum ( L. plantarum ) has been developed for delivery of heterologous oxalate decarboxylase for the prevention of CaOx urolithiasis and the administration of recombinant L . plantarum prevents the enteric hyperoxaluria in experimental rats .…”

Selection of mutants tolerant of oxidative stress from respiratory cultures of Lactobacillus plantarum C17

“…; Sasikumar et al . ), the identification of the mutations would require whole‐genome sequencing of the mutants and of the wild‐type, which is beyond the scope of this work. The mutant C17‐m58 has been selected for further comparative studies (chemostat cultivation in defined composition medium) to clarify some metabolic aspects about respiration in Lact.…”