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C W Tabor

Publications and source records attributed to C W Tabor.

At least 55 records · Page 3Linked to original sources
PubMed1982

Polyamine requirement for efficient translation of amber codons in vivo.

Multiplication of several amber mutants of bacteriophage T7 was decreased in two polyamine-deficient mutants of Escherichia coli K-12 carrying amber suppressors, relative to the multiplication of wild type bacteriophage T7 in the same hosts. In contrast the same T7 amber bacteriophages multiplied well in these strains when supplemented with polyamines. The requirement for polyamines for optimal translation of amber codons in vivo was confirmed by showing that infection of polyamine-depleted E. coli with bacteriophage T7 carrying an amber mutation in gene 1 resulted in an increased accumulation of the amber fragment of the gene 1 protein and a decreased accumulation of the full-length gene 1 protein compared with infection of an amine-supplemented culture. These results indicate that one important function of polyamines in vivo is concerned with protein translation and the protein-synthesizing ribosomal complex.

Codon↗
PubMed1982

The biochemistry, genetics, and regulation of polyamine biosynthesis in Saccharomyces cerevisiae.

We have studied the enzymes and genes involved in the biosynthesis of putrescine, spermidine, and spermine in Saccharomyces cerevisiae. Mutants have been isolated with defects in the biosynthetic pathway as follows: spe10 mutants, deficient in ornithine decarboxylase, cannot make putrescine, spermidine, or spermine; spe2 mutants, lacking S-adenosylmethionine decarboxylase, cannot make spermidine or spermine; spe3 mutants, lacking putrescine aminopropyltransferase, cannot make spermidine or spermine; and spe4 and spe40 mutants, lacking spermidine aminopropyltransferase, contain no spermine and permit growth of spe10 mutants. Studies with these mutants have shown that in yeast: 1) polyamines are absolutely required for growth; 2) putrescine is formed only by decarboxylation or ornithine; 3) two separate aminopropyltransferases are required for spermidine and spermine synthesis; 4) spermine and spermidine are important in the regulation of ornithine decarboxylase and the amines exert this control by a posttranslational modification of the enzyme; and 5) spermidine or spermine is essential for sporulation of yeast and for the maintenance of the double-stranded RNA killer plasmid. Recent studies in amine-deficient mutants of Escherichia coli have shown an important role of the polyamines in protein synthesis in vivo.

Adenosylmethionine Decarboxylase↗
PubMed1981

Ornithine decarboxylase from Saccharomyces cerevisiae. Purification, properties, and regulation of activity.

Ornithine decarboxylase has been purified 1,500-fold to homogeneity from a spe2 mutant of Saccharomyces cerevisiae which lacks S-adenosylmethionine decarboxylase and is derepressed for ornithine decarboxylase. The ornithine decarboxylase is a single polypeptide (Mr = 68,000) and requires a thiol and pyridoxal phosphate for activity. Addition of 10(-4) M spermidine and 10(-4) M spermine to the growth medium reduces the activity of the enzyme by 90% in 4 h. However, immunoprecipitation studies showed that the extracts of polyamine-treated cells contain as much enzyme protein as normal cell extracts. This loss of ornithine decarboxylase activity is probably due to a post-translational modification of enzyme protein because we found no evidence for any inhibitor of activity in the polyamine-treated cells.

Antigen-Antibody Complex↗
PubMed1981

Streptomycin resistance (rpsL) produces an absolute requirement for polyamines for growth of an Escherichia coli strain unable to synthesize putrescine and spermidine [delta(speA-speB) delta specC].

The presence of certain rpsL (strA) mutations in a strain of Escherichia coli that cannot synthesize putrescine or spermidine because of deletions in ornithine decarboxylase, arginine decarboxylase, and agmatine ureohydrolase, converts a partial requirement for polyamines for growth into an absolute requirement.

Drug Resistance, Microbial↗
PubMed1981

Mutants of Saccharomyces cerevisiae deficient in polyamine biosynthesis: studies on the regulation of ornithine decarboxylase.

We have isolated the following mutants in the polyamine biosynthetic pathway in yeast: (i) spe10 mutants, which have no ornithine decarboxylase activity and therefore cannot make putrescine; (ii) spe2 mutants, which have no adenosylmethionine decarboxylase and therefore cannot make spermidine or spermine; (iii) spe3 mutants, which have no putrescine aminopropyltransferase and therefore cannot make spermidine and spermine, and (iv) spe4 and spe40 mutants (suppressors of spe10 mutations), which have no spermidine aminopropyltransferase and therefore cannot make spermine. These mutants show that (i) yeast has an absolute requirement for these amines for growth (ii) putrescine in the absence of spermidine and spermine supports growth at one-sixth the wild type rate; (iii) intracellular spermine controls the ornithine decarboxylase activity and thus mutants that cannot make spermine are derepressed for ornithine decarboxylase; (iv) Saccharomyces cerevisiae can make putrescine only by one pathway, i.e., ornithine decarboxylase; (v) spermidine and spermine are synthesized by different aminopropyltransferases in yeast; and (vi) spermidine and/or spermine are absolutely required for both sporulation and maintenance of the double-stranded RNA "killer" plasmid. We have purified ornithine decarboxylase to homogeneity and shown that loss of ornithine decarboxylase activity resulting from growth with added spermidine and spermine is the result of post-translational modification.

Adenosylmethionine Decarboxylase↗
PubMed1980

S-Adenosylmethionine synthetase from Escherichia coli.

Adenosylmethionine (AdoMet) synthetase has been purified to homogeneity from Escherichia coli. For this purification, a strain of E. coli which was derepressed for AdoMet synthetase and which harbors a plasmid containing the structural gene for AdoMet synthetase was constructed. This strain produces 80-fold more AdoMet synthetase than a wild type E. coli. AdoMet synthetase has a molecular weight of 180,000 and is composed of four identical subunits. In addition to the synthetase reaction, the purified enzyme catalyzes a tripolyphosphatase reaction that is stimulated by AdoMet. Both enzymatic activities require a divalent metal ion and are markedly stimulated by certain monovalent cations. AdoMet synthesis also takes place if adenyl-5'yl imidodiphosphate (AMP-PNP) is substituted for ATP. The imidotriphosphate (PPNP) formed is not hydrolyzed, permitting dissociation of AdoMet formation from tripolyphosphate cleavage. An enzyme complex is formed which contains one equivalent (per subunit) of adenosylmethionine, monovalent cation, imidotriphosphate, and presumably divalent cation(s). The rate of product dissociation from this complex is 3 orders of magnitude slower than the rate of AdoMet formation from ATP. Studies with the phosphorothioate derivatives of ATP (ATP alpha S and ATP beta S) in the presence of Mg2+, Mn2+, or Co2+ indicate that a divalent ion is bound to the nucleotide during the reaction and provide information on the stereochemistry of the metal-nucleotide binding site.

Adenylyl Imidodiphosphate↗
PubMed1980

Regulatory mutations affecting ornithine decarboxylase activity in Saccharomyces cerevisiae.

We isolated several strains of Saccharomyces cerevisiae containing mutations mapping at a single chromosomal gene (spe10); these strains are defective in the decarboxylation of L-ornithine to form putrescine and consequently do not synthesize spermidine and spermine. The growth of one of these mutants was completely eliminated in a polyamine-deficient medium; the growth rate was restored to normal if putrescine, spermidine, or spermine was added. spe10 is not linked to spe2 (adenosylmethionine decarboxylase) or spe3 (putrescine aminopropyltransferase [spermidine synthease]). spe 10 is probably a regulatory gene rather than the structural gene for ornithine decarboxylase, since we isolated two different mutations which bypassed spe10 mutants; these were spe4, an unliked recessive mutation, and spe40, a dominant mutation linked to spe10. Both spe4 and spe40 mutants exhibited a deficiency of spermidine aminopropyltransferase (spermine synthase), but not of putrescine aminopropyltransferase. This suggests that ornithine decarboxylase activity is negatively controlled by the presence of spermidine aminopropyltransferase.

Carboxy-Lyases↗
PubMed1980

Construction of an Escherichia coli strain unable to synthesize putrescine, spermidine, or cadaverine: characterization of two genes controlling lysine decarboxylase.

We have previously described a polyamine-deficient strain of Escherichia coli that contained deletions in speA (arginine decarboxylase), speB (agmatine ureohydrolase), speC (ornithine decarboxylase), and speD (adenosylmethionine decarboxylase). Although this strain completely lacked putrescine and spermidine, it was still able to grow at a slow rate indefinitely on amine-deficient media. However, these cells contained some cadaverine (1,5-diaminopentane). To rule out the possibility that the presence of cadaverine permitted the growth of this strain, we isolated a mutant (cadA) that is deficient in cadaverine biosynthesis, namely, a mutant lacking lysine decarboxylase, and transduced this cadA gene into the delta (speA-speB) delta speC delta D strain. The resultant strain had essentially no cadaverine but showed the same phenotypic characteristics as the parent. Thus, these results confirm our previous findings that the polyamines are not essential for the growth of E. coli or for the replication of bacteriophages T4 and T7. We have mapped the cadA gene at 92 min; the gene order is mel cadA groE ampA purA. A regulatory gene for lysine decarboxylase (cadR) was also obtained and mapped at 46 min; the gene order is his cdd cadR fpk gyrA.

Cadaverine↗
PubMed1979

Mutants of Escherichia coli that do not contain 1,4-diaminobutane (putrescine) or spermidine.

Strains of Escherichia coli K12 have been constructed which do not contain any of the polyamines normally present in a wild type strain, namely, 1,4-diaminobutane (putrescine) and spermidine. This phenotype arises as a consequence of the assembly into these strains of deletion mutations in speA (arginine decarboxylase), speB (agmatine ureohydrolase), speC (ornithine decarboxylase), and speD (adenosylmethionine decarboxylase). The polyamine-deficient strains grow indefinitely in the absence of polyamines but with a growth rate one-third of that found in the presence of polyamines. These strains can act as hosts for bacteriophages T4, T7, and f2, although the latter phage is poorly adsorbed; they can also maintain F' factors, ColE1 and P1 plasmids, and lysogeny by bacteriophage lambda. In contrast, the production of bacteriophage lambda in the absence of polyamines is strikingly decreased (greater than 99%) either after infection of a nonlysogen or after induction of a lysogen. A polyamine-deficient Hfr strain can transfer its chromosome to a recipient at a normal rate, but the number of recombinants observed in a cross is decreased approximately 300-fold. No such effect is observed when only the F- recipient strain in a cross is polyamine deficient.

Amino Acids↗
PubMed1979

An enzymatic method permitting early determination of histocompatibility in mixed lymphocyte culture.

The standard mixed lymphocyte culture assay, which measures the incorporation of [3H]thymidine into DNA, usually requires 5 days. We describe a more rapid assay based on changes in the activity of ornithine decarboxylase. An increase in the activity of ornithine decarboxylase was observed in mixed lymphocyte cultures from genetically defined, major histocompatibility complex (MHC)-nonidentical miniature swine as early as 18 hr after plating. No increase was found in mixed cultures from inbred MHC-identical animals. Similar results were obtained with the enzyme S-adenosylmethionine decarboxylase with the increase in activity starting at about 32 hr. There was a good correlation between the ornithine decarboxylase values at 18 hr and the results of the [3H]thymidine incorporation assay on day 5. Preliminary experiments with human lymphocytes revealed similar results.

Adenosylmethionine Decarboxylase↗
PubMed1978

Escherichia coli mutants completely deficient in adenosylmethionine decarboxylase and in spermidine biosynthesis.

Mutants of Escherichia coli deficient in adenosylmethionine decarboxylase, an enzyme in the biosynthetic pathway for spermidine, were isolated after mutagenesis of E. coli K 12 with N-methyl-N-nitro-N-nitrosoguanidine or with the bacteriophage Mu. The mutated gene, designated speD, is at 2.7 min on the E. coli chromosome map. In several of the mutants resulting from Mu insertion both adenosylmethionine decarboxylase activity and spermidine were undetectable. The absence of spermidine from speD strains proves the essential role of adenosylmethionine decarboxylase in the biosynthetic pathway for spermidine. Despite the complete absence of spermidine, these mutants grew at 75% of the wild type rate.

Adenosylmethionine Decarboxylase↗
PubMed1978

Identification of pyruvate in S-adenosylmethionine decarboxylase from rat liver.

S-Adenosylmethionine decarboxylase has been purified to homogeneity (26,000-fold) from rat liver. The enzyme has a molecular weight of 155,000 and a subunit molecular weight of 42,000. One mole of covalently bound pyruvate was found to be present per mole of enzyme subunit. This is the first mammalian enzyme found to contain covalently linked pyruvate.

Adenosylmethionine Decarboxylase↗
PubMed1978

Isolation and characterization of Saccharomyces cerevisiae mutants deficient in S-adenosylmethionine decarboxylase, spermidine, and spermine.

Four mutants were isolated from Saccharomyces cerevisiae that are deficient in S-adenosylmethionine decarboxylase (spe2). All four mutants are chromosomal and fall into a single complementation group tightly linked to arg1. Since one of the mutants contained a temperature-sensitive activity, this complementation group defines the structural gene. Mutants totally lacking enzymic activity did not contain spermidine or spermine and had a greatly increased doubling time when grown in the absence of these two polyamines. Addition of 10(-6) M spermidine or 10(-5) M spermine, but not putrescine or cadaverine, restored the doubling time to that of the wild type. Diploids formed from a cross of two mutants completely deficient in spermidine and spermine were unable to sporulate in the absence of added spermidine or spermine. We obtained evidence that arg1 was not located on any of the 17 known chromosomes, and therefore we postulate that arg1 and spe2 are located on a new 18th chromosome.

Adenosylmethionine Decarboxylase↗
PubMed1977

Identification of a pyruvoyl residue in S-adenosylmethionine decarboxylase from Saccharomyces cerevisiae.

S-Adenosylmethionine decarboxylase from Saccharomyces cerevisiae has been purified to homogeneity. Acid hydrolysis of NaB3H4-reduced enzyme released 2.2 mol of tritiated lactate per mol of dimeric enzyme, indicating that a pyruvate moiety is present. Inhibition of enzymatic activity by NaBH4 reduction and by carbonyl-binding reagents indicates that this pyruvoyl residue is required for the activity of the enzyme. This is the first example reported of a eukaryotic enzyme containing a covalently linked pyruvoyl residue.

Adenosylmethionine Decarboxylase↗