Biodegradation of polyurethanes by Staphylococcus warneri and by microbial co-culture.

With the increasing production and use of polyurethanes (PUs), it is necessary to develop sustainable techniques for the remediation of plastic pollution. The use of microorganisms capable of biodegrading PUs may be an environmentally desirable solution for controlling these plastic contaminants. To contribute to the discovery of alternatives for the mitigation of plastics in the environment, this study aimed to explore the potential of StaphylococcuswarneriUFV_01.21, isolated from the gut of Galleria mellonellalarvae, for biodegradation of PU in pure culture and microbial co-culture with Serratia liquefaciensL135. S. warneri grew using Impranil® PU as the sole carbon source in pure culture and co-culture. With six days of incubation, the biodegradation of Impranil® in Luria Bertani broth was 96, 88 and 76%, while in minimal medium, it was 58, 54 and 42% for S. warneri, S. liquefaciens, and co-culture, respectively. In addition, S. warneri in pure culture or co-culture was able to biodegrade, adhere and form biofilms on the surfaces of Impranil® disks and poly[4,4'-methylenebis (phenyl isocyanate)-alt-1,4-butanediol/di(propylene glycol)/polycaprolactone] (PCLMDI) films. Scanning electron microscopy also revealed biodegradation by detecting the formation of cracks, furrows, pores, and roughness on the surfaces of inoculated PU, both with pure culture and microbial co-culture. This study is the first to demonstrate the potential of S. warneriin PU biodegradation.

Biodegradation of polyurethanes by Serratia liquefaciens L135 and its polyurethanase: In silico and in vitro analyses.

Polyurethanes (PUs) are found in many everyday products and their disposal leads to environmental accumulation. Therefore, there is an urgent need to develop ecologically sustainable techniques to biodegrade and recycle this recalcitrant polymer and replace traditional methods that form harmful by-products. Serratia liquefaciens L135 secretes a polyurethanase with lipase activity, and this study explores the biodegradation of PUs by this bacterium and its enzyme through in silico and in vitro analyses. PUs monomers and tetramers were constructed in silico and tested with modeled and validated structure of the polyurethanase from S. liquefaciens. The molecular docking showed that all PUs monomers presented favorable interactions with polyurethanase (values of binding energy between -84.75 and -121.71 kcal mol-1), including PU poly[4,4'-methylenebis (phenyl isocyanate)-alt-1,4-butanediol/di (propylene glycol)/polycaprolactone] (PCLMDI). Due to repulsive steric interactions, tetramers showed less favorable interactions (values between 24.26 and -45.50 kcal mol-1). In vitro analyses evaluated the biodegradation of PUs: Impranil® and PCLMDI; this latter showed high binding energy with this polyurethanase in silico. The biodegradation of Impranil® by S. liquefaciens and its partially purified polyurethanase was confirmed in agar by forming a transparent halo. Impranil® disks inoculated with S. liquefaciens and incubated at 30 °C for six days showed rupture of the PU structure, possibly due to the formation of cracks visualized by scanning electron microscopy (SEM). PCLMDI films were also biodegraded by S. liquefaciens after 60 days of incubation, with the formation of pores and cracks visualized by SEM. The biodegradation may have occurred due to the action of polyurethanase produced by this bacterium. This work provides essential information on the potential of S. liquefaciens to biodegrade PUs through in silico analyses combined with in vitro analyses.

Identification and characterization of a polyurethanase with lipase activity from Serratia liquefaciens isolated from cold raw cow's milk.

Lipases are associated with food spoilage and are also used in various biotechnological applications. In this study, we sought to purify, identify, and characterize a lipase from S. liquefaciens isolated from cold raw cow's milk. The lipase partially purified by ultrafiltration and gel filtration showed a specific activity of 2793 U/mg. By zymography, the enzyme presented approximately 65 kDa, and LC-MS/MS allowed the identification of a polyurethanase with a conserved domain of family I.3 lipase. The modeled and validated structure of polyurethanase was able to bind to different fatty acids and urethane by molecular docking. The polyurethanase showed optimum activity at pH 8.0 and 30 °C. In the presence of ions, activity was decreased, except for Ca2+, Mg2+, and Ba2+. Reducing agents did not alter the activity, while amino acid modifiers reduced enzyme activity. It is concluded that polyurethanase with lipase activity represents a potential enzyme for the deterioration of milk and dairy products, as well as a candidate for industrial applications.

Biochemical Characterization of an Extracellular Heat-Stable Protease from Serratia liquefaciens Isolated from Raw Milk.

The protease Ser2 secreted by the psychrotrophic strain Serratia liquefaciens L53, a highly proteolytic strain isolated from Brazilian raw milk was purified and characterized. Using azocasein as substrate, Ser2 exhibited activity in a wide range of pH (5 to 10) and temperature (4 to 60 °C). The optimal activity was detected at pH 8.0 and at a temperature of 37 °C. This protease, still active at 4, 7, and 10 °C, was strongly inhibited by chelating agents and by dithiothreitol, a reducing agent. These results confirmed that Ser2 belongs to the peptidase family M10 and requires Ca2+ , Zn2+ , and disulfide bridges for stability. This protease is able to hydrolyze three kinds of casein in the preferential order of κ→ ß→ α-casein. Highly heat-stable in skimmed, semi-skimmed, and whole milk at 140°C with D-values of 2.8, 3.9, and 4.5 min, respectively, Ser2 showed a residual activity between 87 and 100 percent after heat-treatment of 65 °C for 30 min, 72 °C for 20 s, and 140 °C for 4 s that are commonly used in dairy industries. As the protease AprX that is mainly secreted by Pseudomonas genus, Ser2 could be one of the main causes of UHT milk destabilization during storage.