The enzyme that cleaves apolipoprotein A-II upon in vitro incubation of human plasma high-density lipoprotein-3 with blood polymorphonuclear cells is an elastase

The proteolytic activity directed against apolipoprotein A-II (apo-A-II) which is released from human blood polymorphonuclear cells (PMN) when they are incubated with human plasma high-density lipoprotein-3 (HDL3) was studied to assess the properties and site specificity of the enzyme. When 125I-apo-A-II-labeled HDL3 was incubated with the PMN protease at 37 degrees C, a complete cleavage of apo-A-II was observed which paralleled the formation of bands of approximately 11,000 and 7,000 daltons by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The 7,000-dalton component had the following N-terminal sequence: NH2-Thr-Asp-Tyr-Gly-Lys-Asp-Leu-Met-Glu-Lys. This corresponds to residues 19 through 28 of the intact apo-A-II monomer. Methoxysuccinyl (MeO-Suc)-Ala-Ala-Pro-Val-chloromethylketone-(CH2Cl) caused a 90% inhibition of apo-A-II hydrolysis at the highest concentration tested (6 X 10(-4)M). Besides apo-A-II, the PMN enzyme also hydrolyzed a synthetic substrate, MeO-Suc-Ala-Ala-Pro-Val-4-nitroanilide and its 4-methylcoumaryl-7-amide analogue. The protease appeared to have a mass of 28,000 daltons as assessed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the [3H]diisopropylfluorophosphate-labeled PMN enzyme. That the PMN enzyme which cleaves apo-A-II is an elastase was derived from the following criteria: 1) cleavage at the Val-X bond in apo-A-II and in the two synthetic substrates studied; 2) prevention of the cleavage by MeO-Suc-Ala-Ala-Pro-Val-CH2Cl, a known specific elastase inhibitor; and 3) a mass comparable to that reported for a pure PMN elastase. These studies establish that apolipoproteins can be suitable substrates for enzymes of the elastase family.     

Protonic and cationic changes during cyclical cation transport driven by electron transfer

A method is described for the subcellular fractionation of goldfish xanthophores. The procedure produces relatively pure fractions of caroteniod droplets, pterinosomes, cytosol and what appears to be plasma membrane. The presence of a distinct pattern of proteins is shown to be associated with the carotenoid droplets. Treatment of the xanthophores with ACTH affects the buoyant density of some carotenoid droplets and stimulates the phosphorylation of a polypeptide associated with the carotenoid droplets.     

Analysis of the antigenic and immunogenic properties of protein M of the influenza virus by an immunoenzyme method

A test system permitting the detection of influenza virus protein M at a concentration of 0.1-0.5 ng/ml in ELISA has been developed. The use of this system made it possible to detect influenza viruses A and B directly in crude virus-containing material and clinical samples obtained from influenza patients. During the outbreak of influenza in the spring of 1983 ELISA was successfully used for the rapid diagnosis of influenza, and some of its advantages in comparison with the conventional immunofluorescence test were thus demonstrated. To overcome difficulties arising from the low immunogenic potency of protein M, in the process of obtaining diagnostic sera and ascitic fluids the animals were immunized with the conjugate of protein M and polyelectrolite, which ensured considerable activation of humoral immune response.     

Molecular evidence for the expression of nicotinic acetylcholine receptor alpha-chain in mouse thymus.

The presence and structure of nicotinic acetylcholine receptor (nAChR) in the thymus has been a subject of interest for many years because of its possible role in the pathogenesis of the autoimmune disease myasthenia gravis. Using the polymerase chain reaction with primers specific for the alpha-chain of nAChR (nAChR-alpha), an 880-bp homologous band was found after amplification of cDNA prepared from mouse thymus, thymic medullary and cortical epithelial cell lines, but not from thymocytes or kidney. Sequencing of the polymerase chain reaction product from the thymus and thymic medullary and cortical epithelial lines showed identity with skeletal muscle nAChR-alpha over the region examined. This region includes the domains of the molecule on which B cell and T cell autoantigenic targets have been described. No evidence was found in mouse tissue for the exon 3A, which has been described in human muscle and the human rhabdomyosarcoma cell line TE671. Our results provide evidence at the RNA level for the expression of the nAChR-alpha on stromal cells but not on thymocytes in normal murine thymus and are consistent with a role for intrathymic autoantigen expression in the pathogenesis of myasthenia gravis.     

Identification of N5-methyl-N5-formyl-2,5,6-triamino-4-hydroxypyrimidine as a major adduct in rat liver DNA after treatment with the carcinogens, N,N-dimethylnitrosamine or 1,2-dimethylhydrazine

Human Tamm-Horsfall urinary glycoprotein from an individual of the blood group Sd(a+) phenotype was tritium-labelled by treatment with galactose oxidase and sodium boro[3H]hydride and was then digested with endo-beta-galactosidase. A series of dialysable, labelled fragments was released from which a pentasaccharide was isolated that strongly inhibited the agglutination of Sd(a+) red cells by human anti-Sda serum and hence contained the Sda determinant structure. Reduction, methylation analysis and sequential exo-glycosidase digestion established the structure of the pentasaccharide as: GalNAc beta(1 leads to 4)[NeuAc(2 leads to 3)]Gal beta(1 leads to 4)GlcNAc beta(1 leads to 3)Gal     

Restriction fragment length polymorphisms of horse class II MHC genes observed using various human alpha-and beta-chain cDNA probes

Genomic DNA isolated from 20 horses was digested with up to six restriction endonucleases and subjected to southern blot hybridization analysis using various human class II alpha- and beta-chain cDNA probes. A high degree of restriction fragment length polymorphism (RFLP) was found for the DQ alpha, DP beta, DQ beta and DR beta probes, about 20 polymorphic bands being detected for each. DR alpha showed 2-4 polymorphic bands, whereas no evidence for DP alpha-like genes was found. A number of correlations of RFLPs with individual alloantisera were apparent.     

The activity of jaw elevator muscles during peanut chewing in patients with temporomandibular joint and muscle pain dysfunction syndrome

A synchronized system of EMG and jaw motion tracking device was used to observe some chewing parameters of jaw elevator muscles in 15 patients with temporomandibular joint and muscle pain dysfunction syndrome (TMJ) and 15 normal subjects. Duration of tooth contact (DTC), duration of muscle contraction before tooth contact (DMC), total duration of muscle contraction (DTM) and velocity of jaw movement during peanut chewing were observed. Symptoms of the TMJ patients included pain and tenderness at joints and muscles, and limitation and clicking at joints during jaw movements. It was found that the TMJ patients needed more numerous breaking off strokes before trituration at the occlusal level. There was a longer DMC in the earlier trituration period and TMJ patients had longer DMC than in normals. No difference was found between right and left side chewing or between temporalis and masseter muscles. DTM in the TMJ group was only slightly longer than in normals and the difference between early and late chewing periods was statistically not significant. DTC was only slightly shorter in the TMJ group while the difference between early and late chewing periods in both groups was significant. The average and maximum closing velocities were significantly lower in the TMJ group in both right and left chewing. The difference in the opening phase was not as significant. It was concluded that DMC and jaw closing velocity are more sensitive parameters than DTM and DTC on the diagnosis of TMJ dysfunction with or without occlusal interference. DTM and DTC are parameters more closely related to the influence of occlusal factors.(ABSTRACT TRUNCATED AT 250 WORDS)     

Cyclic AMP is not detectable in Clostridium perfringens

Cyclic AMP was not detected (less than 5 X 10(-9) M intracellular concentration) at any stage of growth or sporulation of two strains of Clostridium perfringens grown with or without methylxanthines. Only Bacillus and Lactobacillus, genera belonging to the same phylogenetic cluster, have previously exhibited undetectable levels of cyclic AMP.     

Comparative greenhouse study of Eucalyptus grandis in vitro plantlets and half-sib seedlings,II. Dry matter accumulation and relative distribution

In vitro directly micropropagated plantlets from three selected five-year-old Eucalyptus grandis Hill ex. Maiden hybrids were compared to their related half-sib seedlings for growth and growth pattern parameters under greenhouse conditions used for operational seedling production. The oven dry weights were determined from stem, leaf, and root samples collected every 40 days for four times. Relative growth rate, net assimilation rates and shoot:root ratio were calculated. Survival was 98% and 95% for plantlets and seedlings, respectively. Significant differences were observed between parents in terms of shoot and root dry weights and their ratios with similar ranking among plantlets and seedlings, suggesting genetic control over these traits. Plantlets started with significantly higher root: shoot ratios and stem, leaf, root, and total dry weight. Although seedlings had higher relative growth and net assimilation rates, all the initial differences decreased sharply over time.