ABSTRACT
Certain members of the Actinobacteria and Proteobacteria are known to degrade polyethylene terephthalate (PET). Here, we describe the first functional PET-active enzymes from the Bacteroidetes phylum. Using a PETase-specific Hidden-Markov-Model- (HMM-) based search algorithm, we identified several PETase candidates from Flavobacteriaceae and Porphyromonadaceae. Among them, two promiscuous and cold-active esterases derived from Aequorivita sp. (PET27) and Kaistella jeonii (PET30) showed depolymerizing activity on polycaprolactone (PCL), amorphous PET foil and on the polyester polyurethane Impranil® DLN. PET27 is a 37.8 kDa enzyme that released an average of 174.4 nmol terephthalic acid (TPA) after 120 h at 30°C from a 7 mg PET foil platelet in a 200 µl reaction volume, 38-times more than PET30 (37.4 kDa) released under the same conditions. The crystal structure of PET30 without its C-terminal Por-domain (PET30ΔPorC) was solved at 2.1 Å and displays high structural similarity to the IsPETase. PET30 shows a Phe-Met-Tyr substrate binding motif, which seems to be a unique feature, as IsPETase, LCC and PET2 all contain Tyr-Met-Trp binding residues, while PET27 possesses a Phe-Met-Trp motif that is identical to Cut190. Microscopic analyses showed that K. jeonii cells are indeed able to bind on and colonize PET surfaces after a few days of incubation. Homologs of PET27 and PET30 were detected in metagenomes, predominantly aquatic habitats, encompassing a wide range of different global climate zones and suggesting a hitherto unknown influence of this bacterial phylum on man-made polymer degradation.
ABSTRACT
The realization that anthropogenic nitrogen (N) deposition is causing significant environmental change in many ecosystems has led to lower emissions of reactive N and deposition rates in many regions. However, the impacts of N deposition on terrestrial ecosystems can be long lasting, with significant inertia in the return of the biota and biogeochemical processes to baseline levels. To better understand patterns of recovery and the factors that may contribute to slow or no responses following declines in N deposition, we followed plant species composition, microbial abundance, N cycling rates, soil pH, and pools of NO3- and extractable cations in an impacted alpine ecosystem following cessation of 12-yr experiment increasing N deposition rates by 0, 20, 40, and 60 kg N·ha-1 ·yr-1 . Simulated N deposition had resulted in a tripling in the cover of the nitrophilic species Carex rupestris, while the dominant sedge Kobresia myosuroides had decreased by more than half at the highest N input level. In addition, nitrification rates were elevated, soil extractable magnesium (Mg2+ ) and pH decreased, and aluminum (Al3+ ) and manganese (Mn2+ ) were elevated at the highest N treatment inputs. Over the nine years following cessation of N additions to the impacted plots, only the cover of the nitrophilic C. rupestris showed any recovery to prior levels. Abundances of both bacteria and fungi were lower with N addition in both treatment and recovery plots. Rates of nitrification and pools of NO3- remained elevated in the recovery plots, likely contributing to the lack of biotic response to the cessation of N inputs. In addition, nutrient base cations (Ca2+ and Mg2+ ) and soil pH remained depressed, and the toxic metal cations (Al3+ and Mn2+ ) remained elevated in recovery plots, also potentially influencing biotic recovery. These results emphasize the importance of considering long-term environmental impacts of N deposition associated with legacy effects, such as elevated N cycling and losses of base cations, in determining environmental standards such as the metrics used for critical loads.
