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The small subunit of iron‐dependent ribonucleotide reductases contains a stable organic free radical, which is essential for enzyme activity and which is localized to a tyrosine residue. Tyrosine‐122 in the B2 subunit of Escherichia coli ribonucleotide reductase has been changed into a phenylalanine. The mutation was introduced with oligonucleotide‐directed mutagenesis in an M13 recombinant and verified by DNA sequencing. Purified native and mutant B2 protein were found to have the same size, iron content and iron‐related absorption spectrum. The sole difference observed is that the mutant protein lacks tyrosyl radical and enzymatic activity. These results identify Tyr122 of E. coli protein B2 as the tyrosyl radical residue. An expression vector was constructed for manipulation and expression of ribonucleotide reductase subunits. It contains the entire nrd operon with its own promoter in a 2.3‐kb fragment from pBR322. Both the B1 and the B2 subunits were expressed at a 25‐35 times higher level as compared to the host strain.

An extensive trial to examine physiological and biochemical changes in perch (Perca fluviatilis) inhabiting coastal waters polluted by bleached kraft mill effluents (BKME) has been carried out. The investigations were performed at four different times of year. Fish from a reference site and from sampling sites 2, 4.5, 8, and 10 km from the discharge point were examined. The results of the investigation show profound effects of BKME on several fundamental biochemical and physiological functions. Typical symptoms in perch from the polluted areas were reduced gonad growth, enlarged liver, and very strong induction of certain cytochrome P-450-dependent enzyme activities in the liver. Elevated levels of ascorbic acid in liver tissue and abnormal carbohydrate metabolism reflect the effluent's ability to cause metabolic disorders. Marked effects on the white blood cell pattern indicate a suppressed immune defence. Alterations in the red blood cell status and in the ion balance suggest that the demand for oxygen by certain tissues was increased and that gill function was impaired, respectively. The toxic effects were most pronounced in fish living up to 4.5 km from the discharge point, but some disturbances (e.g. cytochrome P-450 induction, reduced gonad size, and hematological alterations) were observed in fish caught as far as 8–10 km from the kraft bleach plant.

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