Structure of an antifreeze polypeptide and its precursor from the ocean pout, Macrozoarces americanus

Serum antifreeze polypeptides (AFP) from Newfoundland ocean pout have been resolved by ion exchange chromatography and reverse phase high performance liquid chromatography into at least 12 components. The protein sequences of three of the AFP were determined using a combination of protein Edman degradation and cDNA sequencing. The AFP precursor protein encodes for a preprotein of 87 amino acids with no obvious prosequences. Two of the AFP (SP1-A and SP1-C) were separate gene products with minor amino acid sequence differences. The protein structure of SP1-C precursor is MKSVILTGLLFVLLCVDHMTASQSVVAT QLIPINTALTPAMMEGKVTNPIGIPFAEMSQIVGKQVNTPVAKGQTLMPNMVKTYVAGK. The third AFP (SP1-B) is a post-translation modification product of SP1-C. These experiments indicate that the ocean pout AFP are a multigene family with protein structure different from any other known polypeptide antifreezes.     

Comparative research on erythrocyte anionic adenosine triphosphatase in vertebrates

Studies have been made on the level of activity of anion ATPase and its sensitivity to anions in erythrocytic membranes from fishes, amphibians, reptiles, birds and mammals. Significant variations in these properties of the ATPase were found among the species investigated. Protein composition of erythrocytic membranes was also investigated by means of disc-electrophoresis in polyacrylamide gel in men, rabbit, rat, mouse, hamster, tortoise, crow and starling.     

Metabolic inhibition of size-fractionated marine plankton radiolabeled with amino acids,glucose, bicarbonate,and phosphate in the light and dark

The effects of various metabolic inhibitors (dichlorophenyl dimethylurea, chloramphenicol, cycloheximide, carbonyl cyanide m-chlorophenyl hydrazone) on the accumulation of radiolabeled substrates (amino acids, glucose, bicarbonate, phosphate) by size-fractionated marine microbial plankton from the Sargasso Sea and the eastern Canadian arctic were studied in time-course fashion during experimental incubations either exposed to or shielded from ambient solar radiation. Picoplankton accounted for 65% of the organic substrates and phosphate accumulated by the assemblages. The rate of organic substrate accumulation was stimulated by solar radiation in some cases but inhibited in other cases. Presumably, stimulation and inhibition co-occur and the measured response is the net result arising from these counteracting tendencies. Approximately 40% of H¹⁴ CO₃ ^– accumulation in the Sargasso Sea was associated with the picoplankton. The insensitivity of picoplankton¹⁴C-labeling to cycloheximide suggested active prokaryotic photosynthesis rather than heterotrophic assimilation of¹⁴C-labeled algal photosynthates as the route of labeling. The usefulness of some inhibitors was restricted in this study because of inconsistent correlations between the intended primary metabolic effect and the measured ecological response within the duration of the experiment.     

高等植物叶片中的硝酸还原酶活性稳定因子

从小麦、油菜、浮萍、番茄、烟草的叶片中分离得到NR-SF。不同植物材料中NR及NR-SF能起交叉反应;不同NR-SF影响NR酶动力学性质相同;不同NR-SF的凝胶电泳谱带显示蛋白和糖蛋白性质。NR-SF广泛存在于植物细胞中。     

Somatic embryogenesis and plantlet regeneration in the soybean Glycine max

A tissue culture procedure for the regeneration of somatic embryos and plantlets from somatic cells of the soybean Glycine max is described. Bean pods of soybean cv. TGM119 were immersed in liquid nitrogen for 20 minutes. Young embryos were excised from the immature seeds and cultured to form calli. Calli grown from the young embryos were incubated in liquid culture for two weeks. The liquid suspension culture was filtered to obtain single cells. The soybean cells were cultured for one month in a liquid medium in hanging drop cultures for development into proembryoids. The proembryoids were maintained on a solid growth medium for 40 days. The resultant callus tissue was transferred into MS media containing selected combinations and concentrations of 2,4-Dichlorophenoxyacetic acid, Naphthaleneacetic acid, Kinetin, Benzyladenine and Indoleacetic acid. In the presence of Benzyladenine (0.2 mg/l) and Indoleacetic acid (0.01 mg/l), globular and heart shaped somatic embryos were formed on the surface of the calli. Calli containing somatic embryos were transferred into liquid medium and incubated under low light conditions. After six months further incubation, more than 1,000 plantlets and a large number of somatic embryoids at various developmental stages were obtained per flask.Abbreviations KT kinetin - CM coconut milk - BA benzyladenine - NAA napthalene acetic acid - IAA indole acetic acid - 2,4-D 2,4 dichlorophenoxy acetic acid - MS Murashige and Skoog medium     

T and B cells induce macrophages which suppress proliferation but not lymphokine secretion

In vitro culture of mouse spleen cells for 2 days or more leads to the production of adherent, phagocytic, Thy-1-, Ia+, Lyt-2- cells ("suppressor macrophages") which strongly inhibit the proliferative response of T and B lymphocytes to a variety of stimuli: mitogens, specific antigens, and antigen-nonspecific growth factors. Suppressive activity fails to develop, however, in cultured spleen cells from which nonadherent cells have been removed before the initial 48-hr incubation, and only partial suppression is obtained from cell suspensions from which T cells have been depleted before culture. We find that the requirement for nonadherent cells can be replaced by graded doses of lymphocytes. Lyt-2- and Lyt-2+ T cells are about equally potent in inducing suppressive activity in nonadherent cells. Surprisingly, B cells (containing fewer than 0.1% contaminating T cells) are also able to induce suppression in this system. The suppression induced includes both indomethacin-sensitive and indomethacin-resistant components. Interestingly, not all stages of mitogen-induced T-cell activation are blocked by these adherent cells: proliferation is inhibited, but production of interleukin 2 (IL-2) and interleukin 3 (IL-3) is unaffected.     

Base pairing in wheat germ ribosomal 5S RNA as measured by ultraviolet absorption, circular dichroism, and Fourier-transform infrared spectrometry

Ultraviolet absorption (UV) and circular dichroism (CD) spectra of wheat germ 5S RNA, when compared to tRNAPhe, indicate a largely base-paired and base-stacked helical structure, containing up to 36 base pairs. Fourier-transform infrared (FT-IR) spectra of tRNAPhe and wheat germ ribosomal 5S RNA have been acquired at 30 and 90 degrees C. From the difference of the FT-IR spectra between 90 and 30 degrees C, the number of base pairs in both RNAs was determined by modification of a previously published procedure [Burkey, K. O., Marshall, A. G., & Alben, J. O. (1983) Biochemistry 22, 4223-4229]. The base-pair composition and total base-pair number from FT-IR data are now consistent for the first time with optical (UV, CD, Raman) and NMR results for ribosomal 5S RNA. Without added Mg2+, tRNAPhe gave 18 +/- 2 base pairs [7 A-U and 11 G-C], in good agreement with the number of secondary base pairs from X-ray crystallography [8 A-U, 12 G-C, and 1 G-U]. Within the 10% precision of the FT-IR method, wheat germ 5S RNA exhibits essentially the same number of base pairs [14 A-U, 17 G-C, and 5 G-U; for a total of 36] in the absence of Mg2+ as in the presence of Mg2+ [14 A-U, 18 G-C, and 3 G-U; for a total of 35], in agreement with the UV hyperchromism estimate of G-C/(A-U + G-C) = 0.58.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of physico-chemical factors on anion-sensitive ATPase activity

Effects of temperature, pH and anions on the ATPase activity of submitochondrial particles of rat liver, rat heart, mouse liver, of red blood cell membranes, and of soluble enzyme of rat liver, mouse liver mitochondria were studied. The temperature relationships of membrane-bound and soluble ATPases have the breaks at 18-21 degrees C and 30-32 degrees C. These breaks were not shifted by sulfite, thiocyanate, methanol, glycerol and GTP. The pH changes from 6.0 to 8.5 produced no effect on the temperature relationships of ATPase activities but, strongly influenced the rate of ATPase reaction. The conformity between the obtained data and earlier proposed mechanism of anion control over anion-sensitive ATPase activity was discussed.     

5-Hydroxytryptamine : A modulator of food composition but not quantity?

After a meal of protein, in contrast to a meal of carbohydrate (CHO) at 1915 hr, rats allowed to choose from high carbohydrate and high protein diets during 2000-2100 hr prefer CHO (1). Thus the hypothesis that this regulation of macronutrient selection involves brain 5-hydroxytryptamine (5-HT) metabolism was tested. Compared to three baseline days during which rats (250- 300g ) consumed 1 g CHO, rats fed tryptophan (TRP, 5-HT precursor; 15 mg in 1 g CHO) selected meals higher in protein concentration (35.4% vs 46.6%, F (1,12) = 20.05, p less than 0.001) from 10% and 60% casein diets during 2000-2100 hr. Associated with the higher protein selection was an elevated brain 5-HT turnover in rats killed 30 minutes after consuming CHO + TRP. Pretreating rats with p-chlorophenylalanine, an inhibitor of TRP hydroxylase, blocked this effect of TRP (36.3% vs 37.0%). Fenfluramine (1 and 2 mg/kg i.p. at 1945 hr), which transiently enhances neuronal 5-HT release, increased the rat's relative preference for protein from 28.8% to 37.5% (2 mg/kg, t = 3.21, p less than 0.025) during 2000-2100 hr. These rats, also exhibited a selective preference for CHO between 3-12 hrs post injection which paralleled the known subsequent depletion of 5-HT by fenfluramine. We conclude that the relative proportion of protein and carbohydrate selected in a meal is controlled, at least in part, by prior food effects on brain 5-HT metabolism.     

Mutants of Chinese hamster cells deficient in thymidylate synthetase

Stable mutants of Chinese hamster V79 cells deficient in thymidylate synthetase (TS; E.C. 2.1.1.45) have been selected from cultures grown in medium supplemented with folinic acid, aminopterin, and thymidine (FAT). After chemical mutagenesis, the frequency of colonies resistant to the "FAT" medium increased more than 100-fold over the spontaneous frequency. The optimal expression time of the mutant phenotype was 5-7 days after mutagen treatment. The recovery of FAT-resistant colonies in the selective medium was not affected by the presence of wild-type cells at a density below 9,000 cells per cm2. All 21 mutants tested exhibited thymidine auxotrophy; neither folinic acid nor deoxyuridine could support mutant cell growth. There was no detectable TS activity in all 11 mutants so far examined and only about 50% of wild-type activity in three prototrophic revertants, as measured by whole-cell and cell-free enzyme assays. The apparent Michaelis-Menten constant (Km) for deoxyuridine-5'-monophosphate and inhibition constant (Ki) for 5-fluoro-deoxyuridine-5'-monophosphate, measured by whole-cell enzyme assay, appear to be similar for the wild-type and revertant cell lines. Using 5-fluoro-[6-3H]-2'-deoxyuridine 5'-monophosphate as active site titrant, the relative amounts of TS in crude cell extract from the parental, revertant, and mutant cells were shown to exist in a 1:0.5:0 ratio. Furthermore, the enzymes from two revertants were more heat labile than that of V79 cells. These properties, taken together, suggest that the FAT-resistant, thymidine auxotrophic phenotype may be the result of a structural gene mutation at the TS locus. The availability of such a mutant facilitates studies on thymidylate stress in relation to DNA metabolism, cell growth, and mutagenesis.