title: On the deacetylase activity of Vi bacteriophage III particles.
abstract: 1. Using the complete phage particles as an enzyme, O-acetyl (1 leads to 4)-alpha-D-galacturonan (acetylated pectic acid) as a substrate, and gas-liquid-chromatography for the determination of the acid liberated, the virus-catalysed deacetylation of the polymer was studied. The activity was found to be stable up to about 50 degrees C, and from pH 4.5 to 9, with an optimum at pH 7.8; it was not affected by EDTA, or by 1,10-phenanthroline. The initial reaction velocity (at 37 degrees C) exhibited a simple hyperbolical dependence on the substrate concentration, with Km = 10.5 mM for O-acetyl (independent of virus concentration), and Vmax = 15 nmoles/min and 10(10) plaque forming units. The reaction was, however, rapidly inhibited by a partially deacetylated product (but neither by acetate, nor by pectic acid itself). 2. Using the natural substrate, acetylated (1 leads to 4)-2 amino-2-deoxy-alpha-D-galacturonan (Vi polysaccharide, Vi antigen), and a variety of structural analogues, the following conclusions about the substrate specificity of the Vi phage III deacetylase (acetyl-alpha-1,4-galacturonan acylhydrolase) were reached: (a) acetylated galacturonan is as good a substrate as acetylated aminogalacturonan; (b) of the two substrate diastereomers, acetylated alpha-L-guluronan (also 1 ax leads to 4 ax-linked units, but with axial acetyl residues at C-3), and beta-D-mannuronan (1 eq leads to 4 eq-linkages, and axial acetyl groups at C-2), only the former was acted upon, possibly indicating a specificity for the conformation of the polymer rather than for the configuration of the single residues; (c) all acyl analogues tested, O-monofluoroacetyl, O-propionyl, and O-butyryl galacturonan, were inert, showing a high degree of specificity for O-acetyl; (d) the oligomers, acetylated tri- and digalacturonic acid, as well as methyl-alpha-D-galacturonide, were still deacetylated, although more slowly, demonstrating tolerance of the enzyme of substrate size.
mesh_terms: D000115:Acetylesterase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D012483:Salmonella Phages / Q000201:enzymology* / Q000254:growth & development; D013379:Substrate Specificity; D013696:Temperature
chemical_list: D000115:Acetylesterase