Abstract
The uptake of L-asparagine by Escherichia coli K-12 is characterized by two kinetic components with apparent Km values of 3.5 muM and 80 muM. The 3.5 muM Km system displays a maximum velocity of 1.1 nmol/min per mg of protein, which is a low value when compared with derepressed levels of other amino acid transport systems but is relatively specific for L-asparagine. Compounds providing effective competition for L-asparagine uptake were 4-carbon analogues of the L-isomer with alterations at the beta-amide position, i.e., 5-diazo-4-oxo-L-norvaline (Ki = 4.6 muM), beta-hydroxyamyl-L-aspartic acid (Ki = 10 muM), and L-aspartic acid (Ki = 50 muM). Asparagine uptake is energy dependent and is inhibited by a number of metabolic inhibitors. In a derived strain of E. coli deficient in cytoplasmic asparaginase activity asparagine can be accumulated several-fold above the apparent biosynthetic pool of the amino acid and 100-fold above the external medium. The high affinity system is repressed by culture of cells with L-asparagine supplements in excess of 1 mM and is suggested to be necessary for growth of E. coli asparagine auxotrophs with lower supplement concentrations.
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