Enzymatic transformation of bacterial polyhydroxyalkanoates into repolymerizable oligomers directed towards chemical recycling

The enzymatic transformation into an oligomer was carried out with the objective of developing the chemical recycling of bacterial polyesters. Poly(R-3-hydroxyalkanoate)s (PHAs), such as poly[(R-3-hydroxybutyrate)-co-12%(R-3-hydroxyhexanoate)] and poly[(R-3-hydroxybutyrate)-co-12%(R-3-hydroxyvalerate)], were degraded by granulated Candida antarctica lipase B immobilized on hydrophilic silica (lipase GCA) in a diluted organic solvent at 70 degrees C. The degradation products were cyclic oligomers having a molecular weight of a few hundreds. The obtained cyclic oligomer was readily repolymerized by the same lipase (lipase GCA) to produce the corresponding polyester in a concentrated solution. The cyclic oligomer was copolymerized with epsilon-caprolactone using lipase to produce the corresponding terpolymers having an Mw of 21,000. This is the first example of the enzymatic chemical recycling of bacterial PHAs using lipase. Poly(R-3-hydroxybutyrate) [P(3HB)] was also degraded into the linear-type R-3HB monomer to trimer by P(3HB)-depolymerase (PHBDP) in phosphate buffer at 37 degrees C. The degradation using PHBDP required a longer reaction time compared with the lipase-catalyzed degradation in organic solvent. The monomer composition of the oligomer depended on the origin of the PHBDP. The R-3HB monomer was predominately produced by PHBDP from Pseudomonas stutzeri, while the R-3HB dimer was produced by PHBDP from Alcaligenes faecalis T1. Repolymerization of these oligomers by lipase in concentrated organic solvent produced a relatively low-molecular-weight P(3HB) (e.g., Mw=2,000). Degradation of P(3HB) by lipase in organic solvent into repolymerizable cyclic oligomer and degradation of P(3HB) by PHBDP in buffer into hydroxy acid type R-3HB dimer.     

Fabrication of nanofibers with uniform morphology by self-assembly of designed peptides

Fabrication of controlled peptide nanofibers with homogeneous morphology has been demonstrated. Amphiphilic beta-sheet peptides were designed as sequences of Pro-Lys-X(1)-Lys-X(2)-X(2)-Glu-X(1)-Glu-Pro. X(1) and X(2) were hydrophobic residues selected from Phe, Ile, Val, or Tyr. The peptide FI (X(1)=Phe; X(2)=Ile) self-assemble into straight fibers with 80-120 nm widths and clear edges, as examined by transmission electron microscopy (TEM) and atomic force microscopy (AFM). The fiber formation is performed in a hierarchical manner: beta-sheet peptides form a protofibril, the protofibrils assemble side-by-side to form a ribbon, and the ribbons then coil in a left-handed fashion to make up a straight fiber. These type of fibers are formed from peptides possessing hydrophobic aromatic Phe residue(s). Furthermore, a peptide with Ala residues at both N and C termini does not form fibers (100 nm scale) with clear edges; this causes random aggregation of small pieces of fibers instead. Thus, the combination of unique amphiphilic sequences and terminal Pro residues determine the fiber morphology.     

High-Quality Protein Crystal Growth of Mouse Lipocalin-Type Prostaglandin D Synthase in Microgravity

Lipocalin-type prostaglandin (PG) D synthase (L-PGDS) catalyzes the isomerization of PGH(2) to PGD(2) and is involved in the regulation of pain and of nonrapid eye movement sleep and the differentiation of male genital organs and adipocytes, etc. L-PGDS is secreted into various body fluids and binds various lipophilic compounds with high affinities, acting also as an extracellular transporter. Mouse L-PGDS with a C65A mutation was previously crystallized with citrate or malonate as a precipitant, and the X-ray crystallographic structure was determined at 2.0 ? resolution. To obtain high-quality crystals, we tried, unsuccessfully, to crystallize the C65A mutant in microgravity under the same conditions used in the previous study. After further purifying the protein and changing the precipitant to polyethylene glycol (PEG) 8000, high-quality crystals were grown in microgravity. The precipitant solution was 40% (w/v) PEG 8000, 100 mM sodium chloride, and 100 mM HEPES-NaOH (pH 7.0). Crystals grew on board the International Space Station for 11 weeks in 2007, yielding single crystals of the wild-type L-PGDS and the C65A mutant, both of which diffracted at around 1.0 ? resolution. The crystal quality was markedly improved through the use of a high-viscosity precipitant solution in microgravity, in combination with the use of a highly purified protein.     

Siladenoserinols M–P,sulfonated serinol derivatives from a tunicate

Four new sulfonated serinol derivatives, siladenoserinols M–P (1–4), were isolated from a tunicate of the family Didemnidae collected in Indonesia. Their chemical structures were elucidated by the interpretation of NMR and mass spectroscopic data. Two of them (2 and 4) were revealed to be disulfonate serinol derivatives, and the others were monosulfonates. Siladenoserinols A (5) and B (6), which we previously isolated from the same tunicate, inhibited the p53–Hdm2 interaction with an IC₅₀ value of 2.0?μM. However, 1–4 did not inhibit the activity. The result suggested that the acetyl group in the bicyclic ketal unit and/or the glycerophospholipid moiety in 5 and 6 were responsible for the inhibition of the p53–Hdm2 interaction.