The Mr-50 000 polypeptide of mammalian pyruvate dehydrogenase complex participates in the acetylation reactions

The mammalian pyruvate dehydrogenase complex, Mr 8.5 X 10(6), contains an additional tightly bound 50 000-Mr polypeptide, component X, which copurifies with the intact assembly. Small amounts of the individual E2 and X polypeptides were obtained by elution of the protein bands from SDS/polyacrylamide gels. One-dimensional peptide mapping studies with 125I-labelled lipoyl acetyltransferase (E2) and component X subunits indicate that these two proteins are structurally distinct entities. Similar analysis of purified subunits, initially radiolabelled in the intact complex in the presence of [2-14C]pyruvate and N-ethyl-[2,3-14C]maleimide confirm that distinct 14C-labelled peptides are generated from these two species. These protein-chemical data supplement recent immunological findings, which demonstrate that component X is not a proteolytic fragment of the larger lipoyl acetyltransferase (Mr 70 000) subunit. Incubation of the native PDC in the presence of [2-14C]pyruvate leads to rapid uptake of radiolabel, presumably as acetyl groups, into both E2 and protein X. Specific incorporation of acetyl groups declines to a similar extent on both polypeptides after inhibiting pyruvate dehydrogenase (E1) activity by phosphorylation or omitting thiamine diphosphate (TPP) from the assay mixture. Addition of CoASH promotes the parallel deacetylation of both lipoyl acetyltransferase and protein X in a reaction which displays sensitivity to N-ethylmaleimide.     

A scanning electron microscope study of the effect of an enterotoxin from Clostridium perfringens 8-6 on mice of different ages

Intestinal damage to mice caused by an enterotoxin from a coatless spore mutant of Clostridium perfringens type A (8-6) was examined by scanning electron microscopy. Two distinct types of damage were observed, both of which could be correlated with animal age. Damage appeared to occur in a specific sequence similar to that found in previous studies in rabbits. We conclude that the type of ileal tissue damage reflects the mode of toxin incorporation from the gut, which is a function of animal age.     

Caryospora uptoni n. sp. (Apicomplexa: Eimeriidae) from red-tailed hawks (Buteo jamaicensis borealis)

Oocysts of Caryospora uptoni n. sp. were described from the feces of red-tailed hawks, Buteo jamaicensis borealis. Sporulated oocysts were spherical or subspherical and measured 28.1 by 26.4 micron. The oocyst wall was composed of a yellowish outer layer and brownish inner layer and was about 1.5 micron thick. Neither micropyle, polar granules, nor oocyst residuum were present. A single, spherical sporocyst 18.2 by 17.9 micron was present; a Stieda body was absent. A spherical eccentrically located sporocyst residuum was present in many sporocysts, but it degenerated to form a dispersed granular residuum in other sporocysts. Eight randomly arranged sporozoites, 12.6 by 4.2 micron, were present in each sporocyst; they contained a centrally or slightly posteriorly located nucleus.     

Component X. An immunologically distinct polypeptide associated with mammalian pyruvate dehydrogenase multi-enzyme complex

The mammalian pyruvate dehydrogenase multi-enzyme complex contains a tightly-associated 50 000-Mr polypeptide of unknown function (component X) in addition to its three constituent enzymes, pyruvate dehydrogenase (E1), lipoate acetyltransferase (E2) and lipoamide dehydrogenase (E3) which are jointly responsible for production of CoASAc and NADH. The presence of component X is apparent on sodium dodecyl sulphate/polyacrylamide gel analysis of the complex, performed in Tris-glycine buffers although it co-migrates with the E3 subunit on standard phosphate gels run under denaturing conditions. Refined immunological techniques, employing subunit-specific antisera to individual components of the pyruvate dehydrogenase complex, have demonstrated that protein X is not a proteolytic fragment of E2 (or E3) as suggested previously. In addition, anti-X serum elicits no cross-reaction with either subunit of the intrinsic kinase of the pyruvate dehydrogenase complex. Immune-blotting analysis of SDS extracts of bovine, rat and pig cell lines and derived subcellular fractions have indicated that protein X is a normal cellular component with a specific mitochondrial location. It remains tightly-associated with the 'core' enzyme, E2, on dissociation of the complex at pH 9.5 or by treatment with 0.25 M MgCl2. This polypeptide is not released to any significant extent from E2 by p-hydroxymercuriphenyl sulphonate, a reagent which promotes dissociation of the specific kinase of the complex from the 'core' enzyme. Incubation of the complex with [2-14C]pyruvate in the absence of CoASH promotes the incorporation of radio-label, probably in the form of acetyl groups, into both E2 and component X.     

Changes in density of DNA after interaction with dipicolinic acid and its possible role in spore heat resistance

We examined the effect of interacting dipicolinic acid and its calcium chelate on the wet and dry density of DNA. Complexes are produced whose densities are different from those of the individual components. Also, we observed two modes of binding, one strong the other weak, between DPA or CaDPA and DNA. The strength of the binding modes was reflected in the rate of dissolution of the complexes as monitored by changes in wet density with time and temperature. We conclude from these and other data in the literature that the interaction of dipicolinic acid with DNA not only influences the spore wet density and the ratio of core/core⁺ cortex volume, but may also influence the spore heat resistance.     

Uridine phosphorylase activity among the class mollicutes

Uridine phosphorylase activity has been used to detect mycoplasmas in cell cultures by measuring formation of¹⁴C-uracil from¹⁴C-uridine. In this report we show that all species ofMycoplasma, Acholeplasma, andUreaplasma tested exhibited uridine phorphorylase activity. Among the genusSpiroplasma, serogroups I-1, I-3, I-5, I-7, I-8, IV, XIII, and XIV lacked uridine phosphorylase activity.Present address: Ciba-Geigy, Basel, Switzerland.     

Augmentation of blood flow in delayed random skin flaps in the pig: effect of length of delay period and angiogenesis

Skin capillary blood flow and angiogenesis were studied by radioactive microsphere and morphometry technique, respectively, in delayed random skin flaps in the pig. Skin flaps were delayed for 2, 3, 4, 6, or 14 days. Blood flow was measured 6 hours after complete raising of acute and delayed random skin flaps on the opposite flanks of the same pig. It was observed that the capillary blood flow increased significantly (p less than 0.05) within 2 days of delay compared to the acute skin flaps. This capillary blood flow further increased by about 100 percent between days 2 and 3, started to plateau after day 3, and remained unchanged between days 4 and 14 of delay. This increase in capillary blood flow was mainly in the distal portion of the delayed skin flaps. There was no indication of an increase in the density of arteries in all delay periods studied. Our observations did not support the hypotheses that the delay phenomenon involves angiogenesis or long-term adaptation to ischemia, as have been hypothesized previously. The possible mechanism of delay is discussed.     

Selection of a dye for use in semen transport studies in the cow

A series of experiments was carried out to select a dye and diluent that would facilitate identification of bovine inseminate in the uterus of the live cow when using hysteroscopic techniques. Of the dyes tested, Aniline Blue and the permitted food dye Green S as 1% solutions were the most suitable when added to semen with milk buffer diluent. Neither stained the uterine mucosa nor adversely affected the equipment, and both were visually distinctive during hysteroscopy. Using 1% concentrations of these dyes, sperm motility scores were acceptable before and after slow and fast freezing processes.     

Chemical Nature of Malty Flavor and Aroma Produced by Streptococcus lactis var. maltigenes

Mature skim milk cultures of Streptococcus lactis var. maltigenes were steam distilled at low temperature under reduced pressure. Ethyl ether extracts were prepared from the distillates and analyzed by gas-liquid chromatography and mass spectrometry. Twenty of 31 components detected in the culture distillates were identified positively and 11 tentatively, whereas 10 of 19 components detected in the heated skim milk control were identified positively and 9 tentatively. Among components detected in the culture distillate, but not detected in the heated skim milk distillate, and which have not been previously identified in milk cultures of the organism were phenylacetaldehyde and phenethanol. Quantitative analyses of the volatiles entrained from milk cultures of several strains of S. lactis var. maltigenes revealed a probable relationship between variation in the character of the aroma of the cultures and the alcohol/aldehyde ratio.     

A NEW GENUS OF PROTOSTELIDS SHOWING AFFINITIES WITH CERATIOMYXA

A new genus and species of the Protosteliida (Mycetozoa), Ceratiomyxella tahitiensis, was isolated from dead plant material—var. tahitiensis from Tahiti and var. neotropicalis from Brazil and Colombia. The sporocarps have deciduous spores borne singly on slender hollow stalks; zoocysts with anteriorly flagellate planonts are produced. The trophic stage is comprised of uninucleate to plurinucleate amoeboid cells and reticulate plasmodia; the uninucleate cells become flagellate in water. The prespore cells and spores are plurinucleate. Sexuality has not been demonstrated. Var. tahitiensis has globose spores and produces its zoocysts just after spore germination, whereas var. neotropicalis has subglobose spores and forms zoocysts later in the life cycle. The species is thought to show phylogenetic relationships with Ceratiomyxa, which was recently transferred to the Protosteliida by Olive.