The properties of beta-galactosidases (Escherichia coli) with halogenated tyrosines.
An Escherichia coli tyrosine auxotroph (MR1) with an inducible lacZ was generated by mutagenesis. Of several tyrosine derivatives tested, only m-fluorotyrosine supported the growth of this mutant and allowed synthesis of active beta-galactosidase. The pH profiles of the beta-galactosidase that was obtained when this mutant was grown on m-fluorotyrosine (81.5% of the tyrosine was replaced by m-fluorotyrosine) indicated that a tyrosine may be acting as a general acid-base catalyst and that it (or another tyrosine with the same pKa) may be involved in substrate binding. Inactivation of normal beta-galactosidase by treatment with lactoperoxidase in the presence of I- did not affect affinity-column binding, but incubation of this iodinated beta-galactosidase with chymotrypsin caused a rapid degradation of a portion of the treated enzyme equal to the portion of the activity that was lost. A study with 125I- showed that the rapid degradation was mainly confined to iodinated molecules of enzyme. These studies indicate that iodination of beta-galactosidase does not affect binding ability, but causes the enzyme to lose catalytic activity and become susceptible to chymotryptic action. Chloroperoxidase also caused rapid inactivation of normal beta-galactosidase in the presence of Br- or I-, but there was a lag followed by a slow inactivation in the presence of Cl-.
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Division of Biochemistry, Faculty of Science, University of Calgary, Alta., Canada.
Huber
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D001665:Binding Sites; D002384:Catalysis; D002711:Chloride Peroxidase / Q000378:metabolism; D002918:Chymotrypsin / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D006863:Hydrogen-Ion Concentration; D007544:Isopropyl Thiogalactoside / Q000378:metabolism; D007784:Lactoperoxidase / Q000378:metabolism; D008954:Models, Biological; D007470:Monoiodotyrosine / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D010446:Peptide Fragments; D011487:Protein Conformation; D013329:Structure-Activity Relationship; D014443:Tyrosine / Q000031:analogs & derivatives* / Q000378:metabolism; D001616:beta-Galactosidase / Q000235:genetics / Q000378:metabolism*
D010446:Peptide Fragments; C001245:3-fluorotyrosine; D007544:Isopropyl Thiogalactoside; D014443:Tyrosine; D007784:Lactoperoxidase; D002711:Chloride Peroxidase; D001616:beta-Galactosidase; D002918:Chymotrypsin; D007470:Monoiodotyrosine