In Sweden many advanced sewage treatment plants for BOD and phosphorus removal have to be extended with nitrogen removal. Due to existing plant configuration and wastewater composition, denitrification with supply of an external carbon source can be a cost-effective solution in many cases. At the Klagshamn wastewater treatment plant in Malmo investigations for extensive nitrogen removal have been made in a single-sludge system with pre-precipitation and post-denitrification where methanol was added for denitrification. Results from the tests showed that a high level of nitrogen removal can be reached, and that the process was stable and easy to operate. The process application gave less supplementary cost for an extended nitrogen removal than for upgrading the plant with larger basin volumes. In order to examine the purification performance caused by the addition of methanol, the starting period was followed extensively with online nitrate sensors and daily composite samples. The development of the denitrif ication capacity of the sludge with methanol and acetate as carbon sources was followed and microbiological changes were examined microscopically. Complete denitrification was obtained after approximately one month at 10°C. The denitrification capacity of the sludge with methanol reached that of acetate after about the same time. The microscopic examination revealed a growing population of budding and/or appendaged bacteria, presumably Hyphomicrobium spp, reaching a stable maximum at the time when optimal nitrate removal occurred.

We describe the identification and expression cloning of two novel enzymes, a beta-glucanase and an aspartic protease, secreted from the basidiomycetous yeast Phaffia rhodozyma. A cDNA library from P. rhodozyma CBS 6938 was constructed, and full-length cDNA encoding an endo-1,3(4)-beta-glucanase (bg1) and an aspartic protease (pr1) were cloned by expression cloning in Saccharomyces cerevisiae W3124. The bg1 cDNA encodes a 424-residue precursor protein with a putative signal peptide. The pr1 cDNA encodes a 405-residue prepropolypeptide with an 81-residue leader peptide. The aspartic protease was purified and characterized. It has a molecular mass of 36 kDa, an isoelectric point of pH 7.5, a pH activity optimum at 4.0-6.0, and a temperature activity optimum around 40 degrees C. Both enzymes show only low sequence identity to other known enzymes.

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The ribonucleoside triphosphate, deoxyribonucleoside triphosphate, 3' -diphosphate guanosine 5' -diphosphate (ppGpp), and 5-phosphoribosyl 1-pyrophosphate (PRPP) pools in Escherichia coli B were determined by thin-layer chromatography during changing conditions to ammonium starvation. The intracellular concentrations of all nucleotides were found to change in a well-defined order several minutes before andy observed change in the optical density of the culture. The levels of purine nucleoside triphosphates (adenosine 5' -triphosphate [CTP], dCTP) and uridine nucleotides (uridine 5' -triphosphate, deoxythymidine 5'-triphosphate). The deoxyribonucleotides thus behaved as the ribonucleotides. The levels of ppGpp increased 11-fold after the decrease in uridine nucleotides, when the accumulation of stable ribonucleic acid (RNA) stopped. The level of the nucleotide pool did not stabilize until 30 min after the change in optical density. The pool of dGTP dropped concomitantly with the pool of CTP. The nucleotide precursor PRPP exhibited a transient increase, wtih maximum value of four times the exponential levels at the onset of starvation. Apparently the cell adjusts early to starvation by reducing either the phosphorylating activity or the nucleotide biosynthetic activity. As in other downshift systems, the accumulation of stable RNA stopped before the break in optical density and before the stop in protein accumulation. Cell divisions were quite insensitive to the control mechanisms operating on RNA and protein accumulation under ammonium starvation, since the cells continued to divide for 21 min without any net accumulation of RNA.

The TM1 yeast mutant was transformed with a 2 micron‐derived plasmid (YEp24) which carries a chimaeric gene containing the Escherichia coli chloramphenicol acetyl transferase (CAT) gene fused to the 5′‐ and 3′‐flanking regions of the soybean leghemoglobin (Lb) c3 gene. Expression of the chimaeric CAT gene is controlled specifically by heme at a post‐transcriptional level, most likely by regulating the efficiencies of translation. Expression of another chimaeric gene consisting of the neomycin phosphotransferase (NPTII) gene fused to only the 5′‐flanking region of the Lbc3 gene is regulated by heme in a similar way. Thus, in yeast, heme modulates the translation of the chimaeric mRNAs through interactions with the 5′ Lbc3 non‐coding region.