Production of a recombinant polyester-cleaving hydrolase from Thermobifida fusca in Escherichia coli.

The hydrolase (Thermobifida fusca hydrolase; TfH) from T. fusca was produced in Escherichia coli as fusion protein using the OmpA leader sequence and a His(6) tag. Productivity could be raised more than 100-fold. Both batch and fed-batch cultivations yield comparable cell specific productivities whereas volumetric productivities differ largely. In the fed-batch cultivations final rTfH concentrations of 0.5 g L(-1) could be achieved. In batch cultivations the generated rTfH is translocated to the periplasm wherefrom it is completely released into the extracellular medium. In fed-batch runs most of the produced rTfH remains as soluble protein in the cytoplasm and only a fraction of about 35% is translocated to the periplasm. Migration of periplasmic proteins in the medium is obviously coupled with growth rate and this final transport step possibly plays an important role in product localization and efficacy of the Sec translocation process.

Biodegradation of aliphatic homopolyesters and aliphatic-aromatic copolyesters by anaerobic microorganisms.

The anaerobic degradability of natural and synthetic polyesters is investigated applying microbial consortia (3 sludges, 1 sediment) as well as individual strains isolated for this purpose. In contrast to aerobic conditions, the natural homopolyester polyhydroxybutyrate (PHB) degrades faster than the copolyester poly(hydroxybutyrate-co-hydroxyvalerate) (PHBV). For the synthetic polyester poly(epsilon-caroplacton) (PCL), microbial degradation in the absence of oxygen could be clearly demonstrated; however, the degradation rate is significantly lower than for PHB and PHBV. Other synthetic polyesters such as poly(trimethylene adipate) (SP3/6), poly(tetramethylene adipate) (SP4/6), and aliphatic-aromatic copolyesters from 1,4-butanediol, terephthalic acid, and adipic acid (BTA-copolymers) exhibit only very low anaerobic microbial susceptibility. A copolyester with high amount of terephthalic acid (BTA 40:60) resisted the anaerobic breakdown even under thermophilic conditions and/or when blended with starch. For the synthetic polymers, a number of individual anaerobic strain could be isolated which are able to depolymerize the polymers and selected strains where identified as new species of the genus Clostridium or Propionispora. Their distinguished degradation patterns point to the involvement of different degrading enzymes which are specialized to depolymerize either the natural polyhydroxyalkanoates (e.g., PHB), the synthetic polyester PCL, or other synthetic aliphatic polyesters such as SP3/6. It can be supposed that these enzymes exhibit comparable characteristics as those described to be responsible for aerobic polyester degradation (lipases, cutinases, and PHB-depolymerases).

Propionispora hippei sp. nov., a novel Gram-negative, spore-forming anaerobe that produces propionic acid.

A Gram-negative, spore-forming anaerobe, KS(T), was isolated from an enrichment culture that was set up for anaerobic degradation of the aliphatic polyester poly(propylene adipate). The strain had the cellular organization of Sporomusa, vibrio-shaped cells and terminal round spores, and fermented sugars and sugar alcohols to propionic and acetic acid. Based on the morphological and physiological features as well as on a 16S rRNA gene similarity of 98 %, it was grouped with Propionispora vibrioides. A relatively low DNA-DNA hybridization value with the type strain of this species (47 %), and differences in substrate utilization and spore morphology, suggested that the strain should be classified in a separate species, Propionispora hippei sp. nov., with KS(T) as the type strain (=DSM 15287(T)=ATCC BAA-665(T)).

Maintaining your immune system--one method for enhanced longevity.

The immune system is an important evolutionary invention to battle invaders in young and old organisms. Successful aging in humans who achieve nonagenarian status and beyond depends on how the immune system changes over time. Whether certain immune parameters vary with increased age is influenced by the genotype and lifestyle of the individual.

Production of a polyester degrading extracellular hydrolase from Thermomonospora fusca.

The production of a polyester-degrading hydrolase from the thermophilic actinomycete Thermomonospora fusca was investigated with regard to its potential technical application. Only in the presence of a polyester (random aliphatic-aromatic copolyester from 1,4-butanediol, terephthalic acid, and adipic acid with around 40-50 mol % terephthalic acid in the acid component), the excretion of the extracellular enzyme could be achieved with an optimized synthetic medium using pectin and NH(4)Cl as nitrogen source. Compared to complex media, a significantly higher specific activity at comparable volumetric yields could be obtained, thus reducing the expenditure for purification. The activity profile in the medium is controlled by a complex process involving (1) induction of enzyme excretion, (2) enzyme adsorption on the hydrophobic polyester surface, (3) inhibition of enzyme generation by monomers produced by polyester cleavage, and (4) enzyme denaturation. Diafiltration with cellulose acetate membranes as the sole downstream processing step led to a product of high purity and with sufficient yield (60% of total activity). Scaling-up from shaking flasks to a fermentor scale of 100 L revealed no specific problems. However, the excretion of the hydrolase by the actinomycete turned out to be inhibited by the degradation products (monomers) of the aliphatic-aromatic copolyester used as inductor for the enzyme production. The crude enzyme exhibited generally similar properties (temperature and pH optimum) as the highly purified hydrolase described previously; however, the storage capability and thermal stability is improved when the crude enzyme solution is diafiltrated.

Biodegradable Polymeric Materials-Not the Origin but the Chemical Structure Determines Biodegradability.

It is completely plausible that unmodified materials of natural origin, such as the native macromolecules cellulose or starch, are biodegradable. If these materials are modified then degradation may, depending on the degree of modification, be more difficult or even impossible. In the same manner synthesized macromolecules, whether from renewable or petrochemical sources, could be inert or completey biodegradable, depending on their chemical structure.