Purification and partial characterization of an actin-like protein from cricket early egg plasmodium

Summary The purification of an actin-like protein from cricket egg yolk plasmodia by different selective extraction procedures, ammonium sulphate precipitation, ion exchange and immunoabsorption chromatography is described. Criteria of purity from analytical ultracentrifugation, SDS-disc electrophoresis, and immunoelectrophoresis are presented. Immunodiffusion analysis was used to control the success of the purification procedures.The molecular weight of the monomeric form is 60000±10%. Polymerization to pearl-chain aggregate structures occurs under different conditions in 0.1 M KCl in the presence of ATP. Vinblastine precipitation leads to similar structures. Possibly related structures and the possible rÔle of this protein in organizing movements in the plasmodial system are discussed.     

Mouse-human heterokaryon analysis with a 33258 Hoechst-Giemsa technique

The bibenzimidazol derivative 33258 Hoechst can be used to distinguish microfluorometrically between mouse and human nuclei in heterokaryons. This affords a quick and accurate alternative to autoradiography in the analysis of such heterokaryons. The 33258 Hoechst fluorescence patterns can be converted after irradiation to a Giemsa rendition of the differential staining.     

Proteases of the genus Bacillus. II. Alkaline proteases

The alkaline proteases of B. subtilis NRRL B3411, B. pumilis, and B. licheniformis have been isolated by fractionation followed by ion exchange chromatography and their homogeneity demonstrated. General enzyme properties of the B. sublitis NRRL B3411 alkaline protease have been studied and attempts made to differentiate a group of alkaline proteases. It is clear that the alkaline proteases known as Subtilisins or Subtilopeptidases are not, exclusive to B. subtilis but are common to many Bacilli and therefore the generic name Bacillopeptidases has been proposed. It is clear too that on the basis of the effect of pH on activity, amino acid composition, esterase activity, and immunological cross-reactions the Bacillopeptidases can be divided into two groups or types: (a) Bacillopcptidase A (Subtilisin A or Subtilopeptidase A) which includes Subtilisin Carlsberg, B. licheniformis, and B. pumilis alkaline proteases; ( b ) Bacillopeptidase B (Subtilisin B or Subtilopeptidase B) which includes B subtilis NRRL B3411, Subtilisin Novo, Subtilisin BPN' (Nagarse), alkaline protease Daiwa Kasei, and (probably) B. subtilis var. amylosacchariticus. At present, no further differentiation is possible and whether or not the enzymes within group A or B are identical remains an open question. Methods for examination of crude enzyme mixtures or fermentation beers are described and from the examination of a number of crude enzymes and fermentation beers it appears that organisms producing Bacillopeptidase A do not produce neutral protease or amylase, while organisms producing Bacillopeptidase B produce a neutral protease and amylase as well.     

Isolation, identification, synthesis and biological activity of volatile compounds from the heads of Atta ants

S-(+)-4-methyl-3-heptanone has been identified as the principal alarm pheromone of Atta texana and Atta cephalotes. Both enantiomers of 4-methyl-3-heptanone have been synthesized and their biological activities have been compared on both species of ants. Comparison of the geometric averages of response ratios, at threshold concentration levels on A. texana, showed S-(+)-4-methyl-3-heptanone to be about 100 times more active than the (?) enantiomer. A similar analysis also showed no inhibition of the activity of S-(+)-4-methyl-3-heptanone by the (?) enantiomer. A less rigorous study on A. cephalotes showed S-(+)-4-methyl-3-heptanone to be about 210 times more active than R-(?)-4-methyl-3-heptanone.Both ant species produce 3-octanone, possible trace amounts of 3-octanol, and both diastereomers of 4-methyl-3-heptanol. A. texana also produces (+)-2-heptanol, 2-heptanone, and 3-heptanol. A. cephalotes contains trace amounts of 2-heptanone.     

Tryptophanase in Diverse Bacterial Species

The distribution of tryptophanase was studied. The highest observed specific activity, mumoles per minute per milligram (dry weight) cells, is given in parentheses after each species. Tryptophanase was inducible and repressible in Escherichia coli (.914), Paracolobactrum coliforme (.210), Proteus vulgaris (.146), Aeromonas liquefaciens (.030), Photobacterium harveyi (.035), Sphaerophorus varius (.021), Bacteroides sp. (.048), and Corynebacterium acnes (.042). The enzyme was constitutive and nonrepressible in Bacillus alvei (.013), and was inducible but not repressible by glucose in Micrococcus aerogenes (.036). Indole-positive bacteria were found in fecal or intestinal samples from a variety of animals among the mammals, reptiles, insects, molluscs, fish, crustaceans, and amphibians.     

Graft-vs-host reaction limited to a class II MHC difference results in a selective deficiency in L3T4+ but not in Lyt-2+ T helper cell function

Hybrid mice of the (B6 X bm12)F1 combination were inoculated i.v. with parental B6 spleen cells to induce a class II graft-vs-host disease (GVH). Such mice failed to generate in vitro cytotoxic T lymphocyte (CTL) responses that were dependent upon L3T4+ T helper cell (Th) function (e.g., anti-B6-TNP) but were capable of generating in vitro CTL responses that could be mediated by Lyt-2+ Th cells (anti-allo class I). When Th function was assayed directly by interleukin 2 (IL 2) secretion, class II GVH animals were found to be deficient in L3T4+ but not Lyt-2+ IL 2-secreting Th cells. This selective deficiency in L3T4+ Th function correlates with a selective decrease in class II GVH mice of host-derived derived L3T4+ T cells. In addition, it was found that the spleens of class II GVH mice contained cells capable of selectively suppressing L3T4+ Th function. In contrast, mice in which a class I + II GVH occurred were depleted of both L3T4+ and Lyt-2+ Th function as assessed by IL 2 production. The findings that class II GVH selectively depletes L3T4+ T cells and T cell functions are discussed with respect to the immune function of distinct T cell subsets in normal and diseased states.     

Adrenal and testicular function in 14 patients with adrenoleukodystrophy or adrenomyeloneuropathy

Testicular and adrenal function were evaluated in 12 patients with adrenoleukodystrophy and 2 patients with adrenomyeloneuropathy. Although only 5 subjects had clinical symptoms suggesting adrenal insufficiency, an additional 5 showed laboratory evidence of reduced adrenal reserve. 9 of the 14 patients developed neurological deficits prior to the onset of clinical or biological adrenal insufficiency. In the remaining 5 patients, adrenal insufficiency antedated the appearance of neurological symptoms; 2 of these 5 patients had only laboratory evidence of hypoadrenocorticism, and 3 had both clinical and laboratory abnormalities. None of the prepubertal patients had detectable signs of testicular insufficiency, while 3 of the 7 pubertal/adult patients had elevated serum LH or FSH levels. This mild testicular deficiency was seen only in association with clinical adrenal insufficiency and significant neurological impairment.     

Possible inhibitory function of endogenous 15-hydroperoxyeicosatetraenoic acid on prostacyclin formation in bovine aortic endothelial cells

Arachidonic acid is metabolized via the cyclooxygenase pathway to several potent compounds that regulate important physiological functions in the cardiovascular system. The proaggregatory and vasoconstrictive thromboxane A2 produced by platelets is opposed in vivo by the antiaggregatory and vasodilating activity of prostacyclin (prostaglandin I2) synthesized by blood vessels. Furthermore, arachidonic acid is metabolized by lipoxygenase enzymes to different isomeric hydroxyeicosatetraenoic acids (HETE's). This metabolic pathway of arachidonic acid was studied in detail in endothelial cells obtained from bovine aortae. It was found that this tissue produced 6-ketoprostaglandin F1 alpha as a major cyclooxygenase metabolite of arachidonic acid, whereas prostaglandins F2 alpha and E2 were synthesized only in small amounts. The monohydroxy fatty acids formed were identified as 15-HETE, 5-HETE, 11-HETE and 12-hydroxy-5,8,10-heptadecatrienoic acid (HHT). The latter two compounds were produced by cyclooxygenase activity. Nordihydroguaiaretic acid (NDGA), a rather selective lipoxygenase inhibitor and antioxidant blocked the synthesis of 15- and 5-HETE. It also strongly stimulated the cyclooxygenase pathway, and particularly the formation of prostacyclin. This could indicate that NDGA might exert its effect on prostacyclin levels by preventing the synthesis of 15-hydroperoxyeicosatetraenoic acid (15-HPETE), a potent inhibitor of prostacyclin synthetase. 15-HPETE could therefore act as an endogenous inhibitor of prostacyclin production in the vessel wall.