M. Di Girolamo

Selenalysine and protein synthesis.

Selenalysine is a lysine analog having the gamma-methylene group substituted by a selenium atom. It has been demonstrated that selenalysine is activated and transferred to tRNAlys by either Escherichia coli or rat liver aminoacyl-tRNA synthetases, and inhibits lysine incorporation into polypeptides in protein-synthesizing systems from E. coli, rat liver or rabbit reticulocytes. All tests were performed in comparison with thialysine, a lysine analog having the gamma-methylene group substituted by a sulfur atom. In all the reactions studied, both thialysine and selenalysine act as competitive inhibitors of lysine. With respect to thialysine, selenalysine act as competitive inhibitors of lysine. With respect to thialysine, selenalysine shows a slightly lower activity as lysine inhibitor.

Reutilization of ribosomal protein in vivo for the formation of new ribosomal particles in Escherichia coli B

1. 1. When starved for Mg2+, E. coli B cells almost completely lose the ribosomes. The 105 000 × g supernatant of an extract from Mg2+-starved cells contains many more basic proteins than that from unstarved cells. 2. 2. Mg2+-starved cells, after readdition of Mg2+, are able to synthesize complete ribosomal particles, even in the presence of chloramphenicol. Proteins contained in these particles were synthesized before chloramphenicol was added to the culture. 3. 3. Ribosomal particles synthesized during recovery from Mg2+ starvation in the presence of chloramphenicol contain newly synthesized ribosomal RNA. 4. 4. The possibility of a coordination between ribosomal protein synthesis and ribosomal RNA synthesis is entertained.

β-Selenaproline as competitive inhibitor of proline activation

Abstract β-Selenaproline, a proline analog having the β-methylene group substituted by a selenium atom, has been tested in ATP-PPi exchange reaction catalyzed by either Escherichia coli or rat liver aminoacyl-tRNA synthetases. It has been shown that with both enzymatic systems β-selenaproline does not give rise to ATP-PPi exchange, but specifically inhibits proline activation. The inhibition is of fully competitive type and the K i values, lower than the K m values for proline, show that β-selenaproline binds to the synthetases with high affinity. The inability to form the complex with AMP, taking into account also the behavior of γ-selenaproline and other proline analogs, has been ascribed to the presence of the selenium atom in the β-position.

Properties of ribonucleoprotein particles in chloramphenicol-treated cells of Escherichia coli B

Abstract 1. 1. The electrophoretic pattern of proteins isolated from chloramphenicol particles has been examined by polyacrilamide-gel electrophoresis and compared with that obtained with ribosomal proteins. All proteins isolated from chloramphenicol particles, with the exception of two or three in the upper part of the gel, behave like ribosomal proteins on electrophoresis. 2. 2. The sedimentation characteristics of ribonucleoprotein particles present in the cell before chloramphenicol addition have been examined after the addition of antibiotic to the culture. After 15–30 min of incubation in the presence of chloramphenicol, there is a pronounced decrease in the amount of preexisting 50- and 30-S subunits, and chloramphenicol particles containing RNA synthesized before chloramphenicol addition, appear.

Biochemical characterization of a thialysine-resistant clone of CHO cells.

The intracellular transport and the activation of lysine, thialysine and selenalysine have been investigated in a thialysine-resistant CHO cell mutant strain in comparison with the parental strain. The cationic amino acid transport system responsible for the transport of these 3 amino acids shows no differences between the 2 strains as regards its affinity for each of these amino acids. On the other hand the Vmax of the transport system in the mutant is about double that in the parental strain. The lysyl-tRNA synthetase, assayed both as ATP = PPi exchange reaction and lysyl-tRNA synthesis, shows a lower affinity for thialysine and selenalysine than for lysine in both strains; in the mutant, however, the difference is even greater. Thus the thialysine resistance of the mutant is mainly due to the properties of its lysyl-tRNA synthetase, which shows a greater difference of the affinities for lysine and thialysine with respect to the parental strain.

Thialysine and selenalysine as allosteric inhibitors of E. coli aspartokinase III

SummaryThialysine and selenalysine, two lysine isologs having the γ-methylene group substituted by a sulfur or a selenium atom, respectively, inhibit E. coli lysine-sensitive aspartokinase. The inhibition is specific, reversible and non-competitive. Compared to lysine, the two isologs have a less marked inhibitory effect, but show a similar homotropic cooperativity with a Hills coefficient of about 2. The inhibition by each isolog is additive to that by lysine. Both compounds protect the enzyme against thermal inactivation. Overall, the data reported indicate that thialysine and selenalysine bind to the same allosteric site of lysine, the physiological modulator of the enzyme.

Thialysine, Thiaisoleucine and Thiaproline as Inhibitors in Protein Synthesizing Systems

Thialysine, thiaisoleucine and thiaproline are sulfur-containing analogs of natural aminoacids. Thialysine, or S-2-aminoethyl-cysteine, is a lysine analog having the 4 methylene group substituted by a sulfur atom; thiaisoleucine, or 2-amino-3-methylthio-butyric acid, is an isoleucine analog in which the 4 methylene group of the valerianic acid carbon chain is substituted by a sulfur atom.