PMID:4886277

Citation	Squires, CK and Ingraham, JL (1969) Mutant of Escherichia coli exhibiting a cold-sensitive phenotype for growth on lactose.J. Bacteriol. 97:488-94
Abstract	As part of a study on the effect of low temperature on cellular regulatory processes, a class of lactose-negative mutants of Escherichia coli K-12 was isolated which could use lactose as a sole carbon and energy source at 37 C, but which could not use this sugar at 20 C. The lactose operon of the mutants functioned normally at 20 C. Galactose exhibited a strong inhibitory effect on growth, especially at 20 C. Growth of the mutants on glycerol was stopped at 20 C and slowed considerably at 37 C if galactose was added to the medium. Making the mutants galactose-positive eliminated the cold sensitivity of lactose utilization. One mutant was shown to be galactose-1-phosphate uridyl transferase-negative, galactose-kinase heat-sensitive, and uridine diphosphate-galactose-4-epimerase-positive. It is postulated that the mutant is able to phosphorylate galactose at 20 C (if only at a very low rate), but lacking transferase it is poisoned by the accumulation of galactose-1-phosphate. At 37 C, galactokinase is nonfunctional and the mutant grows on the glucose moiety of lactose.
Links	PubMed
Keywords	Enzyme Induction; Escherichia coli; Galactose; Galactosidases; Genetic Complementation Test; Genetics, Microbial; Glucose; Lactose; Membrane Transport Proteins; Molecular Biology; Mutation; Phosphotransferases; Temperature
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Main Points of the Paper

• Complementation of the gal genes is discussed
  • making the mutant gal⁺ corrects the cold sensitivity of lactose fermentation and overcomes the toxic response to galactose at low temperatures

Methods Used

• Parent strain: KG531-1 (his^-, met^-, thi^-, galT^-, strR, λ^R)
• 2-aminopurine (2-AP) mutagenesis
• β-galactosidase assay
• Galactosidase permease assay (uptake and hydrolysis of ONPG)
• Galactokinase assay
• P1 transductions

Annotations

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Species	Taxon ID	Strain	Gene (if known)	OMP	Phenotype	Details	Evidence	Notes
Escherichia coli	NCBI:562	K-III-4		Temperature-sensitive (conditional) growth on lactose medium Temperature-dependent utilization of carbon source (lactose) Abolished utilization of carbon source (galactose) Temperature-dependent activity of cold-sensitive same as above?	Growth	normal growth rates at 20C on glucose and at 37C on both glucose- and lactose- minimal media	Other	Table 2-Growth curve
Escherichia coli	NCBI:562	K-III-4		Decreased synthesis of beta-galactosidase Decreased number of beta-galactosidase molecules Decreased expression of beta-galactosidase	Metabolic Activity	differential rate of beta-galactosidase synthesis fell to a very low value shortly before the mass had doubled.	Biochemical Assay	Figure 1-LacZ assay vs. dry weight
Escherichia coli	NCBI:562	K-III-4		Abolished utilization of carbon source (galactose) Growth phenotype on galactose medium Toxicity term?	Growth	the addition of galactose alone inhibited the growth of the mutant at both temperatures. galactose-dependent toxicity at 20C and 37C (inhibition of growth)	Other	Figure 2 and Table 6- Growth on galactose vs. lactose vs. glycerol at various temperatures
Escherichia coli	NCBI:562	K-III-4	galK	Temperature-dependent (Conditional) activity of galacto-kinase Temperature-dependent (Conditional) decrease of galactokinase activity	Metabolic Activity	galactokinase-negative at 37C, but positive at 20C	Biochemical Assay	Tables 7 and 8
Escherichia coli	NCBI:562	K-III-4	galT	Temperature-dependent (Conditional) activity of galactose-1-phosphate uridyl transferase Temperature-dependent decrease in galactose-1-phosphate uridyl transferase activity	Metabolic Activity	galactose-1-phosphate uridyl transferase-negative at 20C, but positive at 37C	Biochemical Assay	Table 7- results of complementation tests
Escherichia coli	NCBI:562	KG531-1		Decreased utiliziation of carbon source (galactose) Decreased galactose catabolic process Growth phenotype on galactose medium Decreased growth rate on galactose Toxicity term?	Growth	slight toxicity of galactose when grown at 20C and at 37C reduced growth rate on galactose only	Other	Figure 2-Growth curves comparing lactose, galactose, and glycerol at various temperatures
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