Thermophilic polyester hydrolases (PES-H) have recently
enabled
biocatalytic recycling of the mass-produced synthetic polyester polyethylene
terephthalate (PET), which has found widespread use in the packaging
and textile industries. The growing demand for efficient PET hydrolases
prompted us to solve high-resolution crystal structures of two metagenome-derived
enzymes (PES-H1 and PES-H2) and notably also in complex with various
PET substrate analogues. Structural analyses and computational modeling
using molecular dynamics simulations provided an understanding of
how product inhibition and multiple substrate binding modes influence
key mechanistic steps of enzymatic PET hydrolysis. Key residues involved
in substrate-binding and those identified previously as mutational
hotspots in homologous enzymes were subjected to mutagenesis. At 72
°C, the L92F/Q94Y variant of PES-H1 exhibited 2.3-fold and 3.4-fold
improved hydrolytic activity against amorphous PET films and pretreated
real-world PET waste, respectively. The R204C/S250C variant of PES-H1
had a 6.4 °C higher melting temperature than the wild-type enzyme
but retained similar hydrolytic activity. Under optimal reaction conditions,
the L92F/Q94Y variant of PES-H1 hydrolyzed low-crystallinity PET materials
2.2-fold more efficiently than LCC ICCG, which was previously the
most active PET hydrolase reported in the literature. This property
makes the L92F/Q94Y variant of PES-H1 a good candidate for future
applications in industrial plastic recycling processes.
Summary
There is a dynamic reciprocity between plants and their environment: soil physiochemical properties influence plant morphology and metabolism, and root morphology and exudates shape the environment surrounding roots. Here, we investigate the reproducibility of plant trait changes in response to three growth environments.
We utilized fabricated ecosystem (Eco
FAB
) devices to grow the model grass
Brachypodium distachyon
in three distinct media across four laboratories: phosphate‐sufficient and ‐deficient mineral media allowed assessment of the effects of phosphate starvation, and a complex, sterile soil extract represented a more natural environment with yet uncharacterized effects on plant growth and metabolism.
Tissue weight and phosphate content, total root length, and root tissue and exudate metabolic profiles were consistent across laboratories and distinct between experimental treatments. Plants grown in soil extract were morphologically and metabolically distinct, with root hairs four times longer than with other growth conditions. Further, plants depleted half of the metabolites investigated from the soil extract.
To interact with their environment, plants not only adapt morphology and release complex metabolite mixtures, but also selectively deplete a range of soil‐derived metabolites. The Eco
FAB
s utilized here generated high interlaboratory reproducibility, demonstrating their value in standardized investigations of plant traits.
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