A cascade of sulfur transferases delivers sulfur to the sulfur-oxidizing heterodisulfide reductase-like complex.

A heterodisulfide reductase-like complex (sHdr) and novel lipoate-binding proteins (LbpAs) are central players of a wide-spread pathway of dissimilatory sulfur oxidation. Bioinformatic analysis demonstrate that the cytoplasmic sHdr-LbpA systems are always accompanied by sets of sulfur transferases (DsrE proteins, TusA, and rhodaneses). The exact composition of these sets may vary depending on the organism and sHdr system type. To enable generalizations, we studied model sulfur oxidizers from distant bacterial phyla, that is, Aquificota and Pseudomonadota. DsrE3C of the chemoorganotrophic Alphaproteobacterium Hyphomicrobium denitrificans and DsrE3B from the Gammaproteobacteria Thioalkalivibrio sp. K90mix, an obligate chemolithotroph, and Thiorhodospira sibirica, an obligate photolithotroph, are homotrimers that donate sulfur to TusA. Additionally, the hyphomicrobial rhodanese-like protein Rhd442 exchanges sulfur with both TusA and DsrE3C. The latter is essential for sulfur oxidation in Hm. denitrificans. TusA from Aquifex aeolicus (AqTusA) interacts physiologically with AqDsrE, AqLbpA, and AqsHdr proteins. This is particularly significant as it establishes a direct link between sulfur transferases and the sHdr-LbpA complex that oxidizes sulfane sulfur to sulfite. In vivo, it is unlikely that there is a strict unidirectional transfer between the sulfur-binding enzymes studied. Rather, the sulfur transferases form a network, each with a pool of bound sulfur. Sulfur flux can then be shifted in one direction or the other depending on metabolic requirements. A single pair of sulfur-binding proteins with a preferred transfer direction, such as a DsrE3-type protein towards TusA, may be sufficient to push sulfur into the sink where it is further metabolized or needed.

Carbon Fixation in the Chemolithoautotrophic Bacterium Aquifex aeolicus Involves Two Low-Potential Ferredoxins as Partners of the PFOR and OGOR Enzymes.

Aquifex aeolicus is a microaerophilic hydrogen- and sulfur -oxidizing bacterium that assimilates CO2 via the reverse tricarboxylic acid cycle (rTCA). Key enzymes of this pathway are pyruvate:ferredoxin oxidoreductase (PFOR) and 2-oxoglutarate:ferredoxin oxidoreductase (OGOR), which are responsible, respectively, for the reductive carboxylation of acetyl-CoA to pyruvate and of succinyl-CoA to 2-oxoglutarate, two energetically unfavorable reactions that require a strong reduction potential. We have confirmed, by biochemistry and proteomics, that A. aeolicus possesses a pentameric version of these enzyme complexes ((αßγδε)2) and that they are highly abundant in the cell. In addition, we have purified and characterized, from the soluble fraction of A. aeolicus, two low redox potential and oxygen-stable [4Fe-4S] ferredoxins (Fd6 and Fd7, E0 = -440 and -460 mV, respectively) and shown that they can physically interact and exchange electrons with both PFOR and OGOR, suggesting that they could be the physiological electron donors of the system in vivo. Shotgun proteomics indicated that all the enzymes assumed to be involved in the rTCA cycle are produced in the A. aeolicus cells. A number of additional enzymes, previously suggested to be part of a putative partial Wood-Ljungdahl pathway used for the synthesis of serine and glycine from CO2 were identified by mass spectrometry, but their abundance in the cell seems to be much lower than that of the rTCA cycle. Their possible involvement in carbon assimilation is discussed.

Short-term physiologic response of the green microalga Picochlorum sp. (BPE23) to supra-optimal temperature.

Photobioreactors heat up significantly during the day due to irradiation by sunlight. High temperatures affect cell physiology negatively, causing reduced growth and productivity. To elucidate the microalgal response to stressful supra-optimal temperature, we studied the physiology of Picochlorum sp. (BPE23) after increasing the growth temperature from 30 °C to 42 °C, whereas 38 °C is its optimal growth temperature. Cell growth, cell composition and mRNA expression patterns were regularly analyzed for 120 h after increasing the temperature. The supra-optimal temperature caused cell cycle arrest for 8 h, with concomitant changes in metabolic activity. Accumulation of fatty acids was observed during this period to store unspent energy which was otherwise used for growth. In addition, the microalgae changed their pigment and fatty acid composition. For example, palmitic acid (C16:0) content in the polar fatty acid fraction increased by 30%, hypothetically to reduce membrane fluidity to counteract the effect of increased temperature. After the relief of cell cycle arrest, the metabolic activity of Picochlorum sp. (BPE23) reduced significantly over time. A strong response in gene expression was observed directly after the increase in temperature, which was dampened in the remainder of the experiment. mRNA expression levels associated with pathways associated with genes acting in photosynthesis, carbon fixation, ribosome, citrate cycle, and biosynthesis of metabolites and amino acids were downregulated, whereas the proteasome, autophagy and endocytosis were upregulated.