Chloramphenicol damages bacterial DNA.

L(+)-$\text{threo}$-chloramphenicol induces reversion of His^?$\text{Salmonella typhimurium}$ strains TA100 and TA1535 in the conventional Ames' assay without microsomal activation. Any mutagenicity of D(?)-$\text{threo}$-chloramphenicol was masked by toxicity. Similarly, a sensitive fluctuation test showed mutagenesis with L(+)-$\text{threo}$-chloramphenicol at concentrations of 0.5 μM and above but the D(?) isomer proved to be toxic even at these low levels. The L(+) isomer caused single strand breaks in the DNA of $\text{Escherichia coli}\text{B}\text{r}$ and $\text{Salmonella typhimurium}$ strains TA1535, TA100 and TA1976. The D(?) isomer caused breaks in $\text{Escherichia coli}\text{B}\text{r}$ and $\text{Salmonella typhimurium}$ TA1976 although it was less effective and it did not produce DNA breaks in TA1535 or TA100.     

A nitrosated arginine derivative, a powerful mutagen

An arginine derivative, benzoyl-L-arginineamide, when nitrosated, showed a powerful mutagenic action on $\text{E. coli}$ and $\text{Salmonella typhimurium}$. The active principle was identified to be 4-benzoylamido-4-carboxamidon(N-nitroso)butylcyanamide. The mutagenic activity of the new compound was more than 30 times higher than that of N-methyl-N′-nitro-N-nitrosoguanidine at neutral pH.     

The occurrence of pili associated with a plasmid of the W compatibility group

Using electron microscopy, it was found that the acquisition of the W group drug resistance plasmid S-a by normally pilusless bacterial strains was associated with the appearance of pili. The loss of drug resistance markers in presumed R^? revertants was accompanied by a loss of pili. The numbers of pili present on transconjugant strains of the three bacterial species tested were 3.2 pili/cell for $\text{Salmonella typhimurium}$, and 0.19 pili/cell for both $\text{Escherichia coli}$ and $\text{Pseudomonas aeruginosa}$. Negatively stained pili were about 12 nm thick and varied in length from 23 nm to 3,370 nm.     

Effects of yohimbine and naltrexone in counteraction of the morphine straub tail and the amphetamine-potentiated morphine straub tail in mice

The alpha-adrenergic blocking agent, yohimbine, prevented the production of the morphine Straub tail reaction in mice, although on a mg dose basis it was only about $\text{1}\text{400}$ as potent as the narcotic antagonist naltrexone by subcutaneous injection. Likewise, yohimbine prevented the potentiation of the morphine Straub tail reaction by amphetamine, being about $\text{1}\text{170}$ as active as naltrexone. Preliminary studies with another alpha-adrenergic blocking agent, phentolamine, indicated that it also inhibited the production of the Straub tail reaction by morphine, although it appeared to be somewhat weaker than yohimbine in this respect. These results suggest the involvement of alpha-adrenergic mechanisms in the production of the morphine Straub tail reaction and in the potentiation of the morphine Straub tail reaction by amphetamine.     

High mutagenicity and toxicity of a diol epoxide derived from benzo[a]pyrene

(±)-7β,8α-Dihydroxy-9β,10β-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene (BP 7,8-diol-9,10-epoxide) is a suspected metabolite of benzo[a]pyrene that is highly mutagenic and toxic in several strains of $\text{Salmonella}\text{typhimurium}$ and in cultured Chinese hamster V79 cells. BP 7,8-diol-9,10-epoxide was approximately 5, 10 and 40 times more mutagenic than benzo[a]pyrene 4,5-oxide (BP 4,5-oxide) in strains TA 98 and TA 100 of $\text{S.}\text{typhimurium}$ and in V79 cells, respectively. Both compounds were equally mutagenic to strain TA 1538 and non-mutagenic to strain TA 1535 of $\text{S.}\text{typhimurium}$. The diol epoxide was toxic to the four bacterial strains at 0.5–2.0 nmole/plate, whereas BP 4,5-oxide was nontoxic at these concentrations. In V79 cells, the diol epoxide was about 60-fold more cytotoxic than BP 4,5-oxide.     

Metabolic activation of 3-methylcholanthrene: mutagenic and transforming activities of the 9,10-dihydrodiol.

3-Methylcholanthrene and five related dihydrodiols have been tested for microsome-mediated mutagenicity towards $\text{Salmonella}\text{typhimurium}$ TA100 and for the induction of mutation to 8-azaguanine resistance in V79 Chinese hamster cells and malignant transformation in M2 mouse fibroblasts. In both mutagenicity test systems, the 9,10-diol was considerably more active than either the parent hydrocarbon, the related $\text{cis}$-2α,3-diol, the $\text{trans}$-4,5-, the $\text{trans}$-7,8- or the $\text{trans}$-11,12-dihydrodiols. At a non-toxic concentration (1μg/ml medium), the 9,10-diol induced the formation of more transformed malignant foci in cultures of M2 cells than 3-methylcholanthrene and the other diols were either inactive or only weakly active in this test system. The results obtained indicate that the 9,10-dihydrodiol derived from 3-methylcholanthrene is involved, presumably following conversion into the corresponding vicinal diol-epoxide, 9,10-dihydro-9,10-dihydroxy-3-methylcholanthrene 7,8-oxide, in the metabolic activation of this carcinogenic polycyclic hydrocarbon.     

Identification and property of the mutagenic principle formed from a food-component, methylguanidine, after nitrosation in simulated gastric juice

The active principle responsible for the mutagenicity of methylguanidine nitrosated in weakly acidic conditions and in simulated gastric juice was identified to be methylnitrosocyanamide. This compound was quite similar to N-methyl-N'-nitro-N-nitrosoguanidine in their mutagenic quality, while its mutagenic activity was about 16 times higher for a strain of $\text{Salmonella typhimurium}$ than that of the latter at neutral pH.     

Microsome-mediated mutagenicities of the dihydrodiols of 7,12-dimethylbenz[a]anthracene: high mutagenic activity of the 3,4-dihydrodiol.

7,12-Dimethylbenz[a]anthracene and its 3,4-, 5,6-, 8,9- and 10,11-dihydrodiols have been tested for mutagenicity towards $\text{S}$. $\text{typhimurium}$ TA100 in the presence of rat-liver post-mitochondrial supernatants from Aroclor-treated rats. At non-toxic concentrations, the non-K-region 3,4-dihydrodiol was six-fold more active than the parent hydrocarbon. At these concentrations, the 8,9-dihydrodiol showed some mutagenic activity, but the 5,6- and 10,11-dihydrodiols were inactive.     

Controls of citrate synthase activity

The inhibition of citrate synthase by a variety of nucleotides and polycarboxylate compounds is not unexpected since many of the compounds are substrate analogs of citrate synthase. These effectors are interesting by virtue of the fact that many of them are intermediates and/or end products in the metabolic path of which citrate synthase can be considered the first committed step. As a consequence, it is possible to propose regulation of citrate synthase by ATP (or phosphorylation potential) by acyl CoA (acylation level) and NADH (redox potential). Aside from these putative controls, it is possible that the major control of citrate synthase activity is by changes in the concentration of its substrates acetyl CoA and oxalacetate.I discuss in this review the many factors that must be considered before one can decide whether or not interactions between metabolites and enzymes observed in an $\text{in vitro}$ catalytic situation have metabolic relevance. These factors include 1) the concentrations of substrates at the enzyme site, 2) the concentrations of effectors at the enzyme site, 3) the presence of modifying substances, and 4) the difference in behavior of an enzyme at its concentration $\text{in vivo}$ compared to its concentration $\text{in vitro}$. In the case of citrate synthase as is generally true for other enzymes, no accurate knowledge of these factors are available $\text{in vitro}$ so that little can be said concerning the $\text{in situ}$ control of citrate synthase, which may be the result of all the factors acting in concert. The studies of effectors on enzymes $\text{in vitro}$ can only serve as a guideline for parameters to study when techniques are available to study control of enzymes $\text{in situ}$.     

The NAD-linked α-glycerophosphate dehydrogenase of trypanosomes

NAD-linked α-glycerophosphate dehydrogenase plays a key role in the α-glycerophosphate cycle of $\text{Trypanosoma brucei}$. The activity in cell lysates was ample for this role. The enzyme was activated by salts (e.g. MgCl₂ or NaCl); it had a broad pH-optimum for the reduction of dihydroxyacetone phosphate centred at pH 7.4, with an apparent K_m of 0.5 mM; and it was weakly bound to particulate components of cell lysates. The enzyme from $\text{T. vivax}$ was similar to that of $\text{T. brucei}$. These trypanosomal enzymes resemble that of the trypanosomatid $\text{Crithidia fasciculata}$, but are rather different from the enzymes of mammals, birds and insects.     

Effect of dietary phosphorus on reticuloendothelial clearance of salmonella organisms in endotoxin treated guinea pigs

Guinea pigs fed purified diets that contained either 0.4% or 1.0% of phosphorus were injected intraperitoneally with crude $\text{Salmonella typhimurium}$ endotoxin. Functional activity of the RES was evaluated after endotoxin administration by determining the initial clearance rates of non-viable ³²P-labeled $\text{Salmonella typhimurium}$. Three days after administration of endotoxin the RES clearance rates were approximately $\text{2}\text{1}\text{2}$ times greater in animals fed 1.0% than in those fed 0.4% of dietary phosphorus. After the RES response had risen to a maximum (5–6 days), there was no difference between dietary groups and RES clearance rates remained high throughout an 18-day period. The increased clearance rate induced by endotoxin involved largely the liver RES. Endotoxin administration led to a decrease in blood inorganic phosphorus in both dietary groups but the concentration in those fed the higher phosphorus level remained significantly higher. It was concluded that the protective action of high phosphorus is mediated at least in part by its beneficial effect on RES function.     

Activation of human breast carcinoma collagenase through plasminogen activator

Latent collagenase activity was detected in the media of a well-characterized line of human breast carcinoma cells maintained for over two years in culture. The media also contained sufficient plasminogen activator to convert extrinsically added plasminogen to plasmin which in turn activated the collagenase. During culture of the breast carcinoma in serum-free medium, collagenase activity was maximum on day 12 whereas plasminogen activator activity changed little with time. Using type I collagen as a substrate, the activated breast tumor collagenase produced $\text{3}\text{4}\text{?}\text{1}\text{4}$ fragments consistent with a mammalian collagenase. These findings suggest a pathologic role of plasminogen activator in the activation of latent collagenase during tumor invasion.A number of investigators have postulated that proteases may play a role in tumor invasion (1–5). Collagenase is one such protease which is active at neutral pH and specifically cleaves triple helical collagen into two ($\text{3}\text{4}\text{?}\text{1}\text{4}$ fragments (6). Secretion of collagenase by tumor cells migrating from the primary mass provides an attractive hypothesis for the mechanism of tumor invasion of surrounding host connective tissue—since the local environment would likely be at neutral pH. Consequently, a number of investigators have reported significant levels of collagenase activity in a wide variety of tumors (7–14). Abramson (13) has correlated aggressive $\text{in vivo}$ growth in carcinomas of the head and neck with collagenase activity, and Kuettner et al. (14) have postulated that inhibitors of collagenase may prevent tumors from invading cartilage.Collagenase is produced in both latent and active forms (6). The latent form can be activated with brief protease treatment (15). Since one of the proteases capable of activating collagenase is plasmin (15), the possibility arose that tumor cells could activate collagenase through plasminogen activator. Plasminogen activator secreted by tumor cells (4, 5) could convert plasminogen zymogen to plasmin which would in turn activate latent tumor collagenase. Testing this hypothesis $\text{in vitro}$ was the subject of the present study.Previous studies on collagenase from human carcinoma (7, 13, 14) have suffered from the drawback that contaminating inflammatory cells and fibroblasts may have been the source of the collagenase. Therefore, we have studied collagenase production from cultured human breast carcinoma cells which have been well characterized to be mammary epithelial in origin, malignant in karyotype, and able to grow in nude mice. Production of collagenase from these cells is therefore unequivocally of human carcinoma origin. The time course of latent collagenase and plasminogen activator secretion by these cultured tumor cells was studied following withdrawal of serum. To test whether plasminogen activator was secreted in sufficient amounts to indirectly activate latent collagenase, collagenase activity of the culture media was studied after the extrinsic addition of plasminogen. Finally, to verify that the tumor-secreted collagenase cleaved type I collagen at a single locus, enzyme degradation products were studied by gel electrophoresis.     

Increased de novo phospholipid biosynthesis in the stimulated rat exocrine pancreas

The $\text{hisU1820}$ mutant (TA799) of $\text{Salmonella}\text{,}\text{typhimurium}$ shows a substantial increase in the levels of ppGpp (MSI) and of pppGpp (MSII) during several types of metabolic shifts. Noticeable amounts of ppGp (MSIII) are also present post-carbon/energy source downshifts and temperature up-shifts. The increased levels of these guanosine polyphosphates were observed despite the absence of the expected reduction in RNA synthesis upon a nutritional downshift. We, therefore, suggest that the $\text{hisU}$ mutation causes an increase in the accumulation of MSI and MSII; and that ppGpp alone is not sufficient to promote restriction of RNA synthesis during a nutritional transition.     

Effects of l-cysteine and O-acetyl-l-serine in the synthesis and mutagenicity of azide metabolite

The ability of L-cysteine to inhibit azide-metabolite synthesis and mutagenecity is investigated in Salmonella typhimurium TA1530 and cys E₆ strains. L-cysteine specifically inhibits the synthesis of the mutagenic azide metabolite as other compounds containing SH group did not affect the production of this metabolite. Azide mutagenicity is completely inhibited by L-cysteine at a concentration (5 μmoles/plate) where the metabolite mutagenicity was not affected. $\text{O-}\text{Acetyl-}\text{L}\text{-serine}$ can reverse the L-cysteine mediated inhibition of the metabolite synthesis and thus mutagenicity in the same strains. These results suggest that $\text{O-}\text{acetyl-}\text{L}\text{-serine}$ may be required to synthesize the azide metabolite or its precursor.     

Molecular characterization of a stable Flac plasmid

F$\text{lac}$S is a thermostable extrachromosomal element isolated in $\text{Salmonella typhimurium}$ which is altered in its replication as compared to its precursor F_ts114$\text{lac}$. Sedimentation of both these plasmids in alkaline sucrose gradients has indicated a difference in their sizes. Contour length measurements of open circular plasmid DNA molecules photographed in the electron microscope have revealed the estimated molecular weight of F_ts114$\text{lac}$ to be 81 × 10⁶ daltons while that of F$\text{lac}$S is 109 × 10⁶ daltons. F$\text{lac}$S may carry a segment of $\text{S. typhimurium}$ chromosomal or cryptic plasmid DNA.     

Chromosomal incompatibility in Escherichia,coli K-12: A new explanation for the observed instability of minichromosome inheritance

The minichromosome pWS6 was unstable in $\text{Escherichia}$,$\text{coli}$ K-12 but became stable upon transfer to $\text{Salmonella}$,$\text{typhimurium}$. The instability of pWS6 was restored when pWS6 was brought back to $\text{E.}$,$\text{coli}$, an observation consistent with the proposed phenomena of chromosomal incompatibility.     

Glucose repression of the inducible catabolic pathway for N-acetylglucosamine in yeast.

In order to investigate the mechanism of glucose repression of the N-acetylglucosamine metabolic enzymes in $\text{Candida}\text{albicans}$, an obligatory aerobic yeast, the activities of the following inducible enzymes were assayed: the N-acetylglucosamine uptake, N-acetylglucosamine kinase and glucosamine-6-phosphate deaminase. In the presence of glucose or other sugars e.g. succinate and glycerol, synthesis of these enzymes took place at a normal rate, suggesting that the hexose produces no catabolite repression in this organism. On the contrary, strong inhibition by glucose was observed on the activities of N-acetylglucosamine uptake and deaminase in N-acetylglucosamine-grown cells of $\text{Saccharomyces}\text{cerevisiae}$, a facultative aerobe. From the results, it is concluded that “glucose effect” or catabolite repression is absent in $\text{Candida}\text{albicans}$, a pathogenic strain of yeast.     

Ascaris suum actin: properties and similarities to rabbit actin.

Unlike other enzymes of the aromatic multienzyme system, chorismate synthase and the aromatic complex of $\text{Neurospora crassa}$ were found to bind to a column of cellulose phosphate and to elute at a relatively high concentration of phosphate (～ 0.2 $\text{M}$). The fact that other enzymes with similar ionic properties failed to bind to phosphocellulose suggests that the binding of the former two enzyme systems is due to a specific affinity for phosphate. This conclusion is not only supported by the fact that these same enzymes did not bind to a column of carboxymethyl cellulose, but also is consistent with the nature of the catalytic reactions of the enzymes. Both the shikimate kinase enzyme of the aromatic complex and chorismate synthase would be expected to have active sites which accomodate a phosphate moiety. We anticipate that other enzymes which involve phospho-substrates will also be amendable to this procedure.     

Activities of enzymes that metabolize platelet-activating factor (1-alkyl-2-acetyl-sn-glycero-3-phosphocholine) in neutrophils and eosinophils from humans and the effect of a calcium ionophore

Enzymatic systems in human blood cells are described for the activation and inactivation of a biologically active phospholipid (1-alkyl-2-acetyl-$\text{sn}$-glycero-3-phosphocholine) with hypotensive, platelet-aggregating, and inflammatory properties. The results document the presence of alkyldihydroxyacetone-phosphate synthase (forms the $\text{O}$-alkyl linkage in lipids), 1-alkyl-2-lyso-$\text{sn}$-glycero-3-phosphocholine:acetyl-CoA acetyltransferase (produces the biologically active molecule), and 1-alkyl-2-acetyl-$\text{sn}$-glycero-3-phosphocholine: acetylhydrolase (destroys the biological activity) in human neutrophils and eosinophils. Both the acetyltransferase and acetylhydrolase activities are increased severalfold after treatment of normal neutrophils with ionophore A23187; however, alkyldihydroxyacetone-phosphate synthase activity is not influenced by the ionophore. Eosinophils isolated from patients with eosinophilia have significantly greater activities of all the enzymes studied than the eosinophils isolated from normal individuals. Our results indicate the acetyltransferase responsible for 1-alkyl-2-acetyl-$\text{sn}$-glycero-3-phosphocholine synthesis may serve an important role in human blood cells that release this biologically active phospholipid. Moreover, the acetyltransferase activity was found to be dramatically influenced by calcium flux.