Search

Bacterial microcompartments (BMCs) are organelles that host specific biochemical reactions for both anabolic and catabolic functions. Engineered morphologically diverse BMCs bearing heterologous enzymatic pathways have shown enhanced productivity for commodity chemicals, which makes BMCs an important focus for metabolic engineering. Gaining control of BMC assembly and incorporation of a heterologous enzymatic cargo has yet to be achieved in thermophiles. Herein, we address this by first conducting a detailed bioinformatic analysis of the propanediol utilization ( pdu ) operon in the thermophile Parageobacillus thermoglucosidasius . We then demonstrated, in vivo , the ability to assemble the native BMCs at an elevated temperature of 60 °C. Heterologous expression of Pdu shell proteins from P. thermoglucosidasius in Bacillus subtilis resulted in the assembly of a single tubular BMC with an average length of 1.4 μm; BMCs assembled after a 20 min induction of expression of the shell operons. Moreover, we show that it is possible to target the monomeric superfolder GFP (msfGFP) to the interior of the compartment by fusion of an N-terminal sequence of the propanediol utilization protein (PduP) of at least 24 amino acids. This study establishes the feasibility of constructing cell factories for small molecules in industrially important Bacillus and Geobacillus spp. by heterologous cargo-carrying BMC production and assembly. Additionally, the study provides experimental confirmation that BMCs are produced in thermophilic bacteria, which opens a path for future research on repurposing the native organelles to provide new functionality at elevated temperatures.

Exposure of the respiratory mucosa to oxygen-enriched air contributes to the generation of the lung damage in both adult respiratory distress syndrome and bronchopulmonary dysplasia. Recent work has identified the nasal submucosal gland as the source of diverse molecules important in mucous membrane host defense. We searched for the presence of antioxidant activity in nasal glandular secretions, the absence of which could possibly predispose to oxygen-induced injury. Employing a low molecular weight preparation of nasal secretions (a pooled concentrate passed over a 10,000-dalton molecular sieve), antioxidant activity capable of inhibiting both horseradish peroxidase and Fenton reagent reactions was discovered. The following lines of evidence suggest that submucosal glands are the source of this activity. (1) Antioxidant activity present in resting, baseline nasal washings is significantly increased after cholinergic stimulation either in response to topical methacholine or induced by a gustatory reflex. (2) Application of atropine reduced the antioxidant activity to baseline levels after either of the cholinergic stimuli. (3) Levels of antioxidant activity correlated very closely with the secretion of lactoferrin, a recognized product secreted solely from the serous cell of the submucosal gland. The antioxidant activity is due to novel, previously unrecognized molecules. This activity is found in nasal secretions containing molecules less than 10,000 daltons, is unaffected by N-ethyl maleimide (which inactivates glutathione, another low molecular weight antioxidant), is not associated with the capacity to reduce cytochrome c (as seen with ascorbic acid), and resides in the water soluble pool of secretions (in contrast to vitamin E, another putative antioxidant).(ABSTRACT TRUNCATED AT 250 WORDS)