Allosteric effect of protons and adenosine triphosphate on hemoglobins from aquatic amphibia

Summary The analysis of hemoglobins from two anurans, one semi-aquatic (Leptodactylus labyrinthicus) and the other aquatic (Pipa carvalhoi), showed several components isolated by CM-cellulose chromatography. The two major components (Hb II and Hb III) ofL. labyrinthicus and the major components (Hb III and Hb IV) ofP. carvalhoi possess functional properties as follows: i.P. carvalnoi Hb III and Hb IV andL. labyrinthicus Hb II had very small Bohr effects (–0.05) and a substantial heterotropic effect with polyphosphates. ii.L. labyrinthicus Hb III produced a normal Bohr effect of –0.17, with no influence of polyphosphates.The authors wish to dedicate this paperin memoriam of Prof. Eraldo Antonini who contributed much to the knowledge of hemoglobins     

Structural requirements of a membrane-spanning domain for protein anchoring and cell surface transport

The membrane-spanning domain of the vesicular stomatitis virus glycoprotein (G) contains 20 uncharged and mostly hydrophobic amino acids. We created DNAs specifying G proteins with shortened transmembrane domains, by oligonucleotide-directed mutagenesis. Expression of these DNAs showed that G proteins containing 18, 16, or 14 amino acids of the original transmembrane domain assumed a transmembrane configuration and were transported to the cell surface. G proteins containing only 12 or 8 amino acids of this domain also spanned intracellular membranes, but their transport was blocked within a Golgi-like region in the cell. A G protein completely lacking the membrane-spanning domain accumulated in the endoplasmic reticulum and was secreted slowly. These experiments indicate that the size of the transmembrane domain is critical not only for membrane anchoring, but also for normal cell surface transport.     

Characterization of human interleukin 2 derived from Escherichia coli.

Interleukin 2 isolated from Escherichia coli cells expressing the human interleukin gene has been characterized. The observed properties of the protein have been compared with those properties which can be deduced from the DNA sequence alone and the published properties of natural human interleukin 2. The purified E. coli-derived interleukin 2 is a monomeric protein of Mr 15 000 with a sedimentation velocity of 1.86S. The amino acid composition of the protein and isoelectric point (7.7) are consistent with that part of the translated DNA sequence of the gene corresponding to the mature protein. A single disulphide bridge was identified between Cys-58 and Cys-105. C.d. suggested that interleukin 2 is predominantly alpha-helical in secondary structure. The E. coli-derived protein differed from natural interleukin 2 in the presence of N-terminal methionine and also in the absence of a carbohydrate moiety. Removal of the coding region for the first three amino acids of the natural interleukin 2 protein sequence (Ala-Pro-Thr) by site-specific mutagenesis resulted in a protein with N-terminal serine. The possibility that the specificity of the E. coli ribosomal methionine aminopeptidase may not recognize the sequence NH2-Met-Xaa-Pro is discussed (where Xaa is any amino acid residue).     

Incorporation of a charged amino acid into the membrane-spanning domain blocks cell surface transport but not membrane anchoring of a viral glycoprotein.

The membrane-spanning domain of the vesicular stomatitis virus glycoprotein (G protein) consists of a continuous stretch of 20 uncharged and mostly hydrophobic amino acids. We examined the effects of two mutations which change the amino acid sequence in this domain. These mutations were generated by oligonucleotide-directed mutagenesis of a cDNA clone encoding the G protein, and the altered G proteins were then expressed in animal cells. Replacement of an isoleucine residue in the center of this domain with a strongly polar but uncharged amino acid (glutamine) had no effect on membrane anchoring or transport of the protein to the cell surface. Replacement of this same isoleucine residue with a charged amino acid (arginine) generated a G protein that still spanned intracellular membranes but was not transported efficiently to the cell surface. The protein accumulated in the Golgi region in about 50% of the cells, and about 20% of the cells had detectable protein levels in a punctate pattern on the cell surface. In the remaining cells the protein accumulated in a vesicular pattern throughout the cytoplasm. Models which might explain the abnormal behavior of this protein are discussed.     

Purification of Tissue Forms (Amastigotes) of Trypanosoma cruzi by Immunoaffinity Chromatography1

A rapid and simple method for the purification of amastigotes of Trypanosoma cruzi from spleens of infected mice is described. A protein A-Scpharose 4B immunoadsorbent column bound with antisera to epimastigotes of T. cruzi was used to purify the tissue forms of this parasite. Host cells and debris are not retained, and parasites can be eluted in high yields and purity. Studies of surface glycoproteins and glycolipids of the purified amastigotes with 18 lectins of various specificities revealed the presence on the parasites of receptors for N-acetylglucosamine, N-acetylgalactosamine, D-galactose, and D-mannose binding lectins.     

Human cellular immune response to measles virus polypeptides

Measles virus polypeptides were separated by polyacrylamide gel electrophoresis and electroeluted from gel sections. The antigenicity of the polypeptides was determined by enzyme-linked immunosorbent assays. The ability of these measles virus antigens to stimulate lymphoproliferation was measured in both high- and low-responder individuals. In contrast to the low-responder lymphocytes which did not proliferate when stimulated with measles virus antigens, the high-responder lymphocytes proliferated when challenged with hemagglutinin, nucleocapsid-associated phosphoprotein, nucleocapsid protein, matrix protein, and fusion protein.     

Behavior of triatomines (Hemiptera: Reduviidae) vectors of Chagas' disease. II. Influence of feeding, lighting and time of day on the number of matings, mating speed and duration of copulation of Panstrongylus megistus (Burm, 1835) under laboratory conditions

To determine the influence of feeding, lighting and time of day on the copulating behavior of Panstrongylus megistus, 480 insect pairs were divided into four groups of 120 each and tested in the following respective situations: without food deprivation (F.D.), with five days of F.D., with ten days of F.D., and with 20 days of F.D. The tests were performed between 9:00 a.m. to 12:00 a.m. and 7:00 p.m. to 10:00 p.m., with light (700-1400 lux) and in the dark (1.4-2.8 lux) and behavior was recorded by the time sampling technique. Mating speed (MS) and duration of copulation (DC) were also calculated for each situation. The maximum frequency of copulation was observed after five days of F.D., at night, in the dark (n = 16), and the minimum was observed for recently-fed pairs, at night, with light (n = 4). Males approached females more often than females approached males. MS was lowest in pairs with twenty days of F.D., at night, with light (mean = 23.0 +/- 16.0 minutes), and highest in recently-fed pairs, during the day, with light (mean = 2.9 +/- 2.5 minutes). DC was shortest in recently-fed insects, during the day, in the dark (mean = 23.5 +/- 6.7 minutes), and longest in recently-fed animals, at night, in the dark (mean = 38.3 +/- 6.9 minutes).     

Relationships among negative life events, physiological reactivity, and health symptomatology

College undergraduates classified as high (n = 25) and low (n = 25) on recent life stress participated in an experiment involving a novel laboratory stressor. Heart rate and pulse arrival time (PAT) were measured during baseline, anticipation, testing, and recovery periods of the experiment. The results did not replicate those obtained by Pardine and Napoli in that high and low life stress subjects did not show differential physiological reactions. In addition, regression analyses failed to demonstrate that physiological reactivity moderated the relationship between life stress and subsequent self-reported psychiatric or physical health symptomatology. The present findings demonstrated neither the stress-buffering effects of physiological reactivity nor a relationship between life stress and reactivity when the latter was conceptualized as an outcome.