Reversal of Azaserine by Phenylalanine

Abstract

Kaplan, Reilly, and Stock (1959) have studied the reversal of azaserine by aromatic amino acids and have hypothesized without any direct evidence that these substances prevent the binding or penetration of the antibiotic into the cell. The present paper presents more direct evidence that this hypothesis is correct. It is shown that azaserine acts on nongrowing cells of Escherichia coli strain ML35 in such a way that subsequent growth in azaserine-free medium leads to increases in permeability, lysis, and death. It has been found that phenylalanine reverses only when it is present during the azaserine treatment and has no effect when present during the subsequent growth period. These results are interpreted to mean that phenylalanine prevents the binding or penetration of azaserine into the cell, but plays no direct role in preventing the metabolic disarrangements caused by azaserine. This work has been aided by a strain (ML35) of E. coli which forms 0-galactosidase constitutively at high levels but lacks a permease. This strain is useful in detecting nonspecific effects on permeability.

Azaserine resistance in Escherichia coli: chromosomal location of multiple genes

“…Previously it was reported that azaserine inhibited the growth of E. coli, and that of all the aromatic amino acids, only L-phenylalanine could reverse this growth inhibition (5,17). There was an inverse relationship between the level of azaserine resistance versus the initial rate of uptake of phenylalanine and the efflux of the amino acid from the cell (16).…”

Antagonisms Between Amino Acids in the Growth of Spirodela Polyrhiza Due to Competitive Amino Acid Uptake

“…Thus antagonism has not been observed if the required amino acid was supplied as a peptide (PRESCOTT, PETERS & SNELL 1953;HIRSCH & COHEN 1953). Moreover antagonism has not been observed when the cells were first exposed to the one, then to the other amino acid (MATHIESON & CATCHESIDE 1955;BROCK & BROCK 1961). More directly it has been shown by MANDELSTAM (1956) that the inhibiting effect of diamines on the growth of arg: and lys : mutants from Escherichia coli was correlated with their ability to restrict the accumulation of arginine and lysine.…”

The LIV-I/LS system as a determinant of azaserine sensitivity of Escherichia coli K-12

“…To analyze the basis of azaserine uptake via the LIV-I and LS systems, transport studies was carried out using 14 C-labeled two representative substrates, leucine and phenylalanine, in the absence and presence of the probable inhibitor azaserine (0.3-1.2 mM). Strain YG228 (DaroP DbrnQ D(livJ -yhhK-livKHMGF ) DpheP ) carrying pYG218 (pSC101 replicon bla þ livH þ M þ G þ F þ ), which is deficient in uptake of phenylalanine and branched-amino acids [28], was transformed with a mini-F-derived plasmid carrying either the livJ gene (pYG237) or the livK gene (pYG239) [28], and then subjected to transport assays in the presence of 1-10 lM leucine and 10-100 lM phenylalanine.…”

“…In earlier works, it was demonstrated that the inhibitory effect of azaserine on E. coli is alleviated effectively in the presence of aromatic amino acids [13], especially phenylalanine [14,15], and partially in the presence of branched-chain amino acids [13]. The alleviating effect of aromatic amino acids on azaserine toxicity could be explained by competition for the uptake of substrates by AroP (a general aromatic amino acid permease with equal affinity to each aromatic amino acid) because all azaserine-resistant mutants isolated so far have lesions in the aroP gene [16].…”

“…Strain YG228 [DaroP DbrnQ D(livJ -yhhK-livKHMGF ) DpheP ] carrying pYG218 (pSC101 replicon bla þ livH þ M þ G þ F þ ) (membranecomponents) was transformed with either pYG237 (Mini-F replicon kan þ livJ þ ) (LIV-I) or pYG239 (Mini-F replicon kan þ livK þ ) (LS)[28], and then used for assaying. Uptake of14 C-labeled L L -leucine [Leu] (1-10 lM) (a and b) and 14 C-labeled L L -phenylalanine [Phe] (10-100 lM) (c and d) by the LIV-I (a and c) and LS (b and d) systems was measured in the absence (Ã) or presence of azaserine (AS) (s, 0.3 mM and M, 1.2 mM).…”