Isolation of unselected mutants of alkaline phosphatase in Escherichia coli through nitrosoguanidine comutation and comparison with natural variants.
We have devised a general procedure to isolate enzymatic variants without selecting or screening for related phenotypic peculiarities of the organism. A high mutation rate at phoA, the structural gene for alkaline phosphatase, is found among N-methyl-N'-nitro-N-nitrosoguanidine-induced proC revertants of Escherichia coli. About 1.6% of such revertants lack alkaline phosphatase, and many others exhibit altered enzyme parameters. Three mutants studied in detail had full enzyme activity but differed from the wild type in electrophoretic mobility, thermostability, and, in one case, optimum pH for enzyme activity. Four other phosphatase variants were discovered in a survey of 50 natural E. coli isolates; their electrophoretic mobility and thermostability were different from those of the wild type. Natural and induced enzyme variants are similar enough to suggest the absence of strong selective pressures in natural populations.
D000469:Alkaline Phosphatase / Q000235:genetics* / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics*; D006358:Hot Temperature; D007527:Isoenzymes / Q000235:genetics; D007700:Kinetics; D009154:Mutation; D009604:Nitrosoguanidines
D007527:Isoenzymes; D009604:Nitrosoguanidines; D000469:Alkaline Phosphatase