The primary structure and physical chemical properties were determined of a nuclease expressed and secreted by Escherichia colt. The plasmid p403-SD2 carried a DNA sequence isolated from Serratia marcescens encoding the enzyme. During cultivation of the E. colt cells, 85% of the enzyme was released to the growth medium. The enzyme was purified and exhibited a single band with a molecular weight about 30,600 daltons on SDS-PAGE similar to nuclease isolated from S. marcescens. The amino acid composition and the amino acid sequence determined directly confirmed the primary structure of 245 amino acids predicted from the DNA sequence, and, in addition, the two disulfide bridges were assigned. Several physical chemical properties were examined. The ability of the enzyme to cross the outer membrane is proposed to depend upon the formation of the proper structures during the folding process.

This structure was established by automated Edman degradation of peptides derived from cyanogen bromide fragmentation and enzymatic digestion with S. aureus V8 protease and chymotrypsin. High pressure liquid chromatography was used to separate the peptides from the different cleavage reactions. The cyanogen fragment residues 2-33 contained the inhibitory activity.

A sensitive assay for acid proteinases, based on tritium labelled hemoglobin, has been used to demonstrate nanogram quantities of acid proteinase in unpasteurized commercial beers. The proteinase was released from brewers' yeast into beer, and the level varied both with 1he yeast strain used and with other process parameters. Acid proteinase was isolated from brewers' yeast and added to beer in amounts similar to the highest activities found in beer samples. This caused a significant reduction in beer foam stability.

A Serratia marcescens nuclease was analyzed by capillary eleCtrOphOred8 under stacking conditions. The purified nuclease sample was obtained after fermentation of the €$-cherichia coil strain MT 102 carrying the plasmid p403-SD2 encoding the S. mnudeam At increasing appiled voltages several peaks emerged in the electropherograms in addltkn to the nuclease peak. The same Mect was obtained by increasing the ratio of running buffer to sample zone specific condllctivlty. Estbnath of the sample zone and running buffer temperature8 suggested that the additional peak8 emerged due to thermal degradation of the nuclease molec u b at temperatures above ca. 40 O C . Thls supposition was supported by experiments In the thermostated range 30-60 OC at experlmental conditions where Joule heating of the sample zone In the capillary could be neglected. I n general, care should be taken when temperature-labile analytes are analyzed by capillary electt'OphOresl8 under stacking conditkns or under normtacking conditions with high Ionic strength buffer8 at high electric field strengths.

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