A cellular protein binds to a conserved sequence in the adenovirus type 2 enhancer.

A sensitive gel retention assay has been utilized to detect proteins from uninfected Hela nuclei which interact with the adenovirus type 2 enhancer. This assay has been employed to monitor fractionation of nuclear extracts. Three enhancer binding factors were resolved by chromatography on DEAE-Sepharose and one of the factors was further purified by chromatography on heparin-Sepharose. DNase protection experiments have shown that the heparin-Sepharose fraction contains a factor which binds predominantly to the conserved sequence GTGGAAATTT present at position 160 in the adenovirus type 2 genome and found in many viral and cellular enhancers. Protection of this sequence from DNase I digestion was abolished by competition with a synthetic duplex oligonucleotide spanning bases 144-181. This region corresponds to the sequence defined by Hen et al. as possessing enhancer function. Competition experiments indicated that the enhancer binding factor also bound, albeit with reduced affinity, to multiple sites in the Ela upstream region located between positions 192 and 353. Within the sequences which compete are regions with homology to the high affinity site at position 160. The enhancer binding factor also binds with high affinity to sequences within the SV40 enhancer demonstrating that this factor interacts with sequences common to both the adenovirus and SV40 enhancers.     

The cysteine proteinases of the pineapple plant.

The pineapple plant (Ananas comosus) was shown to contain at least four distinct cysteine proteinases, which were purified by a procedure involving active-site-directed affinity chromatography. The major proteinase present in extracts of plant stem was stem bromelain, whilst fruit bromelain was the major proteinase in the fruit. Two additional cysteine proteinases were detected only in the stem: these were ananain and a previously undescribed enzyme that we have called comosain. Stem bromelain, fruit bromelain and ananain were shown to be immunologically distinct. Enzymic characterization revealed differences in both substrate-specificities and inhibition profiles. A study of the cysteine proteinase derived from the related bromeliad Bromelia pinguin (pinguinain) indicated that in many respects it was similar to fruit bromelain, although it was found to be immunologically distinct.     

广东哺乳类及两栖类寄生吸虫四新种的记述（吸虫纲：蹲茎科,蛙蠕科）

作者在广东省梅县市发现四个吸虫新种:1.鼠丽色吸虫,新种Reesella ratta sp. nov., 2.后囊蛙蠕吸虫,新种Batrachotrema optstosacca sp. nov., 3.梅县后平睾吸虫,新种Opisthioparorchis meixianensis sp. nov., 4.大卵后平睾吸虫,新种Opisthioparorchis megaloonos sp. nov.,它们分别归隶于蹲茎科Cathaemasiidae及蛙蠕科Bstrachotrematidae。模式标本保存于中山医科大学寄生虫学研究所。     

Affinity purification and biochemical characterization of histolysin, the major cysteine proteinase of Entamoeba histolytica.

Insulin receptor was co-purified from human placenta together with insulin-stimulated kinase activity that phosphorylates the insulin receptor on serine residues. By using this 'in vitro' system, the mechanism of activation of the serine kinase by insulin was explored. Peptide 1150, histone, poly(Glu-Tyr), eliminating Mn2+ (Mg2+ only), treatment at 37 degrees C (1 h), N-ethylmaleimide, phosphate, beta-glycerol phosphate and anti-phosphotyrosine antibody all inhibited insulin-receptor tyrosine kinase activity and the ability of insulin to stimulate phosphorylation of the insulin receptor on serine. Additionally, direct stimulation of the receptor tyrosine kinase by vanadate increased serine phosphorylation of the insulin receptor. Insulin-stimulated tyrosine phosphorylation preceded insulin-stimulated serine phosphorylation of the insulin receptor. The activity of the insulin-sensitive receptor serine kinase was not augmented by cyclic AMP, cyclic GMP, Ca2+, Ca2+ + calmodulin, Ca2+ + phosphatidylserine + diolein or spermine, or inhibited appreciably by heparin. Additionally, the serine kinase phosphorylated casein or phosvitin poorly and was active with Mn2+. This indicates that it is distinct from Ca2+, Ca2+/phospholipid, Ca2+/calmodulin, cyclic AMP- and cyclic GMP-dependent protein kinases, casein kinases I and II and insulin-activated ribosomal S6 kinase. Taken together, these data indicate that a novel species of serine kinase catalyses the insulin-dependent phosphorylation of the insulin receptor and that activation of this receptor serine kinase by insulin requires an active insulin-receptor tyrosine kinase.     

Effects of corticotropin releasing hormone on appetitive behaviors.

Corticotropin releasing hormone (CRH) is a 41-residue hypothalamic neuropeptide that has been shown to have potent behavioral effects in animals and has been implicated in clinical disorders in man. This review focuses on those aspects of the behavioral effects of CRH related to food-associated behaviors. The effects of CRH on food intake are compared with its effects on performances maintained by food presentation, and contrasted with the effects of CRH on performances maintained by other events. The effects of CRH antagonists and drugs that interact with the behavioral effects of CRH are also reviewed, particularly with respect to their direct effects on food intake. Lastly, data assessing the effects of CRH administration on central neurotransmitter levels are presented and compared with levels seen in clinical populations. The effect of CRH on food intake seen in animals is consistent with a putative role for CRH in clinical syndromes where appetite suppression is apparent. Since some of the effects of CRH on food intake are subject to pharmacological intervention, strategies directed at peptidergic mechanisms of psychiatric disorders should be explored.     

Effect of substrate on Ca2(+)-concentration required for activity of the Ca2(+)-dependent proteinases, mu- and m-calpain

The Ca2+ concentrations required for half-maximal activity of mu- and m-calpain purified from bovine skeletal muscle were tested using four different protein substrates and three different synthetic peptide substrates. Hammersten casein, the commonly used substrate for measuring mu- and m-calpain activity, required 2.5 microM Ca2+ for half-maximal activity of mu-calpain and 290 microM Ca2+ for half-maximal activity of m-calpain. When Hammersten casein was dialyzed against 8 M urea and 10 mM EDTA to remove all endogenous Ca2+, it required 1.9 and 290 microM Ca2+ for half-maximal activity of mu- and m-calpain, respectively. Rabbit skeletal muscle myofibrils and rabbit skeletal muscle troponin required 65 microM and 24 microM Ca2+ for half-maximal activity of mu-calpain and 380 microM and 580 microM Ca2+ for half-maximal activity of m-calpain, respectively. The three synthetic substrates tested, Suc-Leu-Tyr-MCA, Boc-Leu-Thr-Arg-MCA, and Suc-Leu-Leu-Val-Tyr-MCA, required 1.6 microM to 3.7 microM Ca2+ for half-maximal activity of mu-calpain and 200 to 560 microM Ca2+ for half-maximal activity of m-calpain.     

Induction by ionizing radiation of the gadd45 gene in cultured human cells: lack of mediation by protein kinase C.

The effect of ionizing radiation on the expression of two DNA-damage-inducible genes, designated gadd45 and gadd153, was examined in cultured human cells. These genes have previously been shown to be strongly and coordinately induced by UV radiation and alkylating agents in human and hamster cells. We found that the gadd45 but not the gadd153 gene is strongly induced by X rays in human cells. The level of gadd45 mRNA increased rapidly after X rays at doses as low as 2 Gy. After 20 Gy of X rays, gadd45 induction, as measured by increased amounts of mRNA, was similar to that produced by the most effective dose of the alkylating agent methyl methanesulfonate. No induction was seen after treatment of either human or hamster cells with 12-O-tetradecanoylphorbol-13-acetate, a known activator of protein kinase C (PKC). Therefore, gadd45 represents the only known mammalian X-ray-responsive gene whose induction is not mediated by PKC. However, induction was blocked by the protein kinase inhibitor H7, indicating that induction is mediated by some other kinase(s). Sequence analysis of human and hamster cDNA clones demonstrated that this gene has been highly conserved and encodes a novel 165-amino-acid polypeptide which is 96% identical in the two species. This gene was localized to the short arm of human chromosome 1 between p12 and p34. When induction in lymphoblast lines from four normal individuals was compared with that in lines from four patients with ataxia telangiectasia, induction by X rays of gadd45 mRNA was less in the cell lines from this cancer-prone radiosensitive disorder. Our results provide evidence for the existence of an X-ray stress response in human cells which is independent of PKC and which is abnormal in taxia telangiectasia.     

Enzyme-linked immunosorbent assays for detecting antibodies to Shiga-like toxin I,Shiga-like toxin II,andEscherichia coli O157:H7 lipopolysaccharide in human serum

Shiga-like toxin-producingEscherichia coli O157:H7 are important causes of bloody diarrhea and hemolytic uremic syndrome. To facilitate the epidemiologic study of these organisms, we developed enzyme-linked immunosorbent assays (ELISAs) for antibodies to Shiga-like toxin I (SLT I), Shiga-like toxin II (SLT II), andE. coli O157 lipopolysaccharide (LPS). We tested serum samples from 83 patients in two outbreaks ofE. coli O157:H7 diarrhea and from 66 well persons. Forty-three patients (52%) had at least one serum sample positive for anti-O157 LPS antibodies; among 26 culture-confirmed patients, 24 (92%) had at least one positive serum sample. Two (3%) of 66 control sera had positive anti-O157 LPS titers. ELISA results for SLT I and II were compared with those of HeLa cell cytotoxicity neutralization assays on both patient and control sera. Neutralization assays detected anti-SLT I antibodies in at least one serum sample from each of 17 (20%) patients and 7 (10.6%) controls, while 16 (19%) patients and 7 controls had positive titers by anti-SLT I ELISA. Although all serum samples, including control sera, showed nonspecific neutralization of SLT II, no antibody titers to SLT II were detected by either neutralization or ELISA. These results indicate that ELISAs for SLT I and SLT II antibodies are comparable to HeLa cell cytotoxicity neutralization assays. Both the ELISAs and neutralization assays are insensitive in detecting infected patients. However, the ELISA for antibodies toE. coli O157 LPS is both sensitive and specific, and may be more useful than assays for antitoxic antibodies in detecting persons withE. coli O157:H7 infection.