The Study of Escherichia coli Proteases. Intracellular Serine Protease of E. coli - an Analogue of Bacillus Proteases

Abstract

Two serine proteases in extracts of Escherichia coli grown to stationary phase were purified to homogeneity using affinity chromatography on gramicidin S-Sepharose 4B. One enzyme was closely related to, if not identical with, the ' trypsin-like' protease I1 of E. coli. The other was capable of cleaving the subtilisin chromogenic substrate N-carbobenzoxy-L-alanyl-Lalanyl-L-leucine-p-nitroanilide and resembled the intracellular serine proteases of Bacillus spp. The amino acid composition of this E. coli protease was similar to that of the Bacillus lichenformis enzyme. These data indicate a relationship between proteolytic enzymes of evolutionary distant Gram-negative Enterobacteriaceae and Gram-positive spore-forming Bacillus.

Isolation and characterization of proteases from Bacteroides melaninogenicus

“…Symbols: 0, protease I; 0, protease II. (6,22,24,28,29,31), but proteases I and II cannot be classified into this category, since neither of these enzymes is inactivated by diisopropylfluorophosphate. Inactivation of proteases I and II by the thiol compounds was marked; this was especially true of protease II, which was inactivated completely by DTT.…”

“…Also, Strongin et al have purified two serine proteases from E. coli by using affinity chromatography (28). One of these enzymes had properties similar to those of protease II of Pacaud and Richard (22), and the other resembled the intracellular protease of Bacillus subtilis (27,29).…”

Purification and Characterization of Protease So, a Cytoplasmic Serine Protease in Escherichia coli

“…When such reagents were used, the control reaction mixture contained equal amounts of the solvent only. L-tyrosine ethyl ester) (28), protease II (N-benzoyl-DL-arginine-p-nitroanilide and N-benzoyl-L-arginine ethyl ester) (27), or a subtilisin-like enzyme called ISP-L-ECO (N-carbobenzoxy-Lalanine-L-alanine-L-leucine-p-nitroanilide) (35).…”

“…The auto a peptide was prepared by autoclaving E. coli j-galactosidase (24). The hydrolysis of N-acetyl-L-phenylalanine-13-naphthyl ester was measured at 328 nm (28), that of N-benzoyl-L-tyrosine ethyl ester was measured at 255 nm (18), those of Nbenzoyl-DL-arginine-p-nitroanilide and N-carbobenzoxy-L-alanine-L-alanine-L-leucine-p-nitroanilide were measured at 410 nm (10,35), and that of N-benzoyl-Larginine-ethyl ester was measured at 255 nm (18). The size distribution of the acid-soluble material produced by protease So was determined by incubating the enzyme (3.0 ,ug) with 35 .g of [3H]casein or 8 ,ug of [14C]globin in a volume of 0.5 ml at 37°C for 1 h.…”

General Properties of a New Ribosomal Aryl-peptidyl Amidase inLactobacillus casei

“…Enzyme activities hydrolyzing succinyl-LPhe-p-nitroanilide type or analogous substrates were systematically considered as proteases· when demonstrated in other bacteria8~21, 22) or even in animal tissues. 23 ) This conclusion is occasionally too hasty, as shown by the present results, and only a detailed study of enzyme specificity on a wide range of substrates can lead to a valid affirmation.…”