Echovirus 22 is an atypical enterovirus

Although echovirus 22 (EV22) is classified as an enterovirus in the family Picornaviridae, it is atypical of the enterovirus paradigm, typified by the polioviruses and the coxsackie B viruses. cDNA reverse transcribed from coxsackievirus B3 (CVB3) RNA does not hybridize to genomic RNA of EV22, and conversely, cDNA made to EV22 does not hybridize to CVB3 genomic RNA or to molecular clones of CVB3 or poliovirus type 1. EV22 cDNA does not hybridize to viral RNA of encephalomyocarditis virus or to a molecular clone of Theiler's murine encephalomyelitis virus, members of the cardiovirus genus. The genomic RNA of EV22 cannot be detected by the polymerase chain reaction using generic enteroviral primers. EV22 does not shut off host cell protein synthesis, and the RNA of EV22 is efficiently translated in vitro in rabbit reticulocyte lysates. Murine enterovirus-immune T cells recognize and proliferate against EV22 as an antigen in vitro, demonstrating that EV22 shares an epitope(s) common to enteroviruses but not found among other picornaviruses.     

Plasma lipoproteins and apolipoproteins in the preruminant calf, Bos spp: density distribution, physicochemical properties, and the in vivo evaluation of the contribution of the liver to lipoprotein homeostasis

The in vivo role of the liver in lipoprotein homeostasis in the preruminant calf, a functional monogastric, has been evaluated. To this end, the hydrodynamic and physicochemical properties, density distribution, apolipoprotein content, and flow rates of the various lipoprotein particle species were determined in the hepatic afferent (portal vein and hepatic artery) and efferent (hepatic vein) vessels in fasting, 3-week-old male preruminant calves. Plasma lipoprotein profiles were established by physicochemical analyses of a series of subfractions isolated by isopycnic density gradient ultracentrifugation. Triglyceride-rich very low density lipoproteins (VLDL) (d less than 1.018 g/ml) were minor plasma constituents (approximately 1% or less of total d less than 1.180 g/ml lipoproteins). The major apolipoproteins of VLDL were apoB-like species, while the complement of minor components included bovine apoA-I and apoC-like peptides. Particles with diameters (193-207 A) typical of low density lipoproteins (LDL) were present over the density interval 1.026-1.076 g/ml; however, only LDL of d 1.026-1.046 g/ml were present as a unique and homogeneous size subspecies, containing the two apoB-like species as major protein components in addition to elevated cholesteryl ester contents. LDL represented approximately 10% of total d less than 1.180 g/ml lipoproteins in fasting plasma from all three hepatic vessels. Overlap in the density distribution of particles with the diameters of LDL and of high density lipoproteins (HDL) occurred in the density range from 1.046 to 1.076 g/ml; these HDL particles were 130-150 A in diameter. HDL were the major plasma particles (approximately 90% of total d less than 1.180 g/ml substances) and presented as two distinct populations which we have termed light (HDLL) and heavy (HDLH) HDL. Light HDL (d 1.060-1.091 g/ml) ranged in size from 120 to 140 A, and were distinguished by their high cholesteryl ester (29-33%) and low triglyceride (1-3%) contents; apoA-I was the principal apolipoprotein. Small amounts of apolipoproteins with Mr less than 60,000, including apoC-like peptides, were also present. Heavy HDL (d 1.091-1.180 g/ml) accounted for almost half (47%) of total calf HDL, and like HDLL, were also enriched in cholesteryl ester and apoA-I; they ranged in size from 93 to 120 A. The protein moiety of HDLH was distinct in its possession of an apoA-IV-like protein (Mr 42,000). Blood flow rates were determined by electromagnetic flowmetry, thereby permitting determination of net lipoprotein balance across the liver. VLDL were efficiently removed during passage through the liver (net uptake 1.06 mg/min per kg body weight).(ABSTRACT TRUNCATED AT 400 WORDS)     

Subunit structure of the galactose and N-acetyl-D-galactosamine-inhibitable adherence lectin of Entamoeba histolytica

The galactose and N-acetyl-D-galactosamine-inhibitable adherence lectin of Entamoeba histolytica is a cell surface protein which mediates parasite adherence to human colonic mucus, colonic epithelial cells, and other target cells. The amebic lectin was purified in 100-micrograms quantities from detergent-solubilized trophozoites by monoclonal antibody affinity chromatography. The adherence lectin was purified 500-fold as judged by radioimmunoassay. The nonreduced lectin had a molecular mass of 260 kDa on sodium dodecyl sulfate-polyacrylamide gel electrophoresis and an isoelectric point of pH 6.2. The amebic lectin reduced with beta-mercaptoethanol consisted of 170- and 35-kDa subunits. Both subunits could be labeled on the cell surface with 125I, and both were metabolically labeled with [3H]glucosamine. The amino termini of the subunits had unique amino acid sequences, and polyclonal antisera to the heavy subunit did not cross-react with the light subunit. The yield of phenylthiohydantoin derivatives from the second and third positions in the sequence of the heavy and light subunits gave a molar ratio of one 170- to one 35-kDa subunit. Antibodies directed to the heavy subunit inhibited amebic adherence to Chinese hamster ovary cells by 100%, suggesting that the heavy subunit is predominantly responsible for mediating amebic adherence.     

Chemotherapy of experimental visceral leishmaniasis in the opossum

Visceral leishmaniasis is a severe chronic disease of people and animals. The disease is caused by several subspecies of a protozoal organism, Leishmania donovani. If not treated, visceral leishmaniasis is often fatal. The most commonly used chemotherapeutic agents to treat the disease are pentavalent antimonials, which can be toxic, must be administered by parenteral routes, and are sometimes ineffective. In this study, meglumine antimoniate, a pentavalent antimony, was compared with WR 6026, an 8-aminoquinoline derivative, as to antileishmanial efficacy. The results indicate that either of these 2 drugs are effective in the suppression of amastigotes in the liver and spleen of the opossum. Despite the marked parasite suppression in the liver and spleen of the infected opossums, the experimental disease was fatal in all of the infected opossums, regardless of the therapy.     

A recombination event that redefines the Huntington disease region

We report both a recombination event that places the Huntington disease gene proximal to the marker D4S98 and an extended linkage-disequilibrium study that uses this marker and confirms the existence of disequilibrium between it and the HD locus. We also report the cloning of other sequences in the region around D4S98, including a new polymorphic marker R10 and conserved sequences that identify a gene in the region of interest.     

1H and 51V NMR studies of the interaction of vanadate and 2-vanadio-3-phosphoglycerate with phosphoglycerate mutase.

The formation of complexes of vanadate with 2-phosphoglycerate and 3-phosphoglycerate have been studied using 51V nuclear magnetic resonance spectroscopy. Signals attributed to two 2,3-diphosphoglycerate analogues, 2-vanadio-3-phosphoglycerate and 2-phospho-3-vanadioglycerate, were detected but were not fully resolved from signals of inorganic vanadate and the anhydride formed between vanadate and the phosphate ester moieties of the individual phosphoglycerates. Equilibrium constants for formation of the two 2,3-bisphosphate analogues were estimated as 2.5 M-1 for 2-vanadio-3-phosphoglycerate and 0.2 M-1 for 2-phospho-3-vanadioglycerate. The results of the binding study are fully consistent with non-cooperativity in the binding of vanadiophosphoglycerate to the two active sites of phosphoglycerate mutase (PGM). 2-Vanadio-3-phosphoglycerate was found to bind to the dephospho form of phosphoglycerate mutase with a dissociation constant of about 1 x 10(-11) M at pH 7 and 7 x 10(-11) M at pH 8. Three signals attributed to histidine residues were observed in the 1H NMR spectrum of phosphoglycerate mutase. Two of these signals and also an additional signal, tentatively attributed to a tryptophan, underwent a chemical shift change when the vanadiophosphoglycerate complex was bound to the enzyme. The results obtained here are in accord with these vanadate-phosphoglycerate complexes being much more potent inhibitors of phosphoglycerate mutase than either monomeric or dimeric vanadate. The dissociation constant of 10(-11) M for 2-vanadio-3-phosphoglycerate is about 4 orders of magnitude smaller than the Km for PGM, a result in accordance with the vanadiophosphoglycerates being transition state analogues for the phosphorylation of PGM by 2,3-diphosphoglycerate.(ABSTRACT TRUNCATED AT 250 WORDS)     

The optional E. coli prr locus encodes a latent form of phage T4-induced anticodon nuclease.

The optional Escherichia coli prr locus restricts phage T4 mutants lacking polynucleotide kinase or RNA ligase. Underlying this restriction is the specific manifestation of the T4-induced anticodon nuclease, an enzyme which triggers the cleavage-ligation of the host tRNALys. We report here the molecular cloning, nucleotide sequence and mutational analysis of prr-associated DNA. The results indicate that prr encodes a latent form of anticodon nuclease consisting of a core enzyme and cognate masking agents. They suggest that the T4-encoded factors of anticodon nuclease counteract the prr-encoded masking agents, thus activating the latent enzyme. The encoding of a tRNA cleavage-ligation pathway by two separate genetic systems which cohabitate E. coli may provide a clue to the evolution of RNA splicing mechanisms mediated by proteins.     

Ethanol transport in Zymomonas mobilis measured by using in vivo nuclear magnetic resonance spin transfer.

For the first time, unidirectional rate constants of ethanol diffusion through the lipid membrane of a microorganism, the bacterium Zymomonas mobilis, were determined, thus replacing indirect inferences with direct kinetic data. The rate constants k1 (in to out) were 6.8 +/- 0.4s(-1) at 29 degrees C and 2.7 +/- 0.3s(-1) at 20 degrees C. They were determined by using 1H selective nuclear magnetic resonance spin magnetization transfer. The measurements were done on l-ml cell suspensions. No addition of radiotracers, withdrawing of aliquots, physical separation methods, or chemical manipulations were required. Until now, the rate constants of ethanol transport in microorganisms have been unknown because ethanol diffuses through the cytoplasmic membrane too quickly for radiolabel approaches. Net velocities of ethanol exchange were calculated from unidirectional rate constants and cytoplasmic volume, which was also determined with the same nuclear magnetic resonance experiments. The results (i) confirmed that ethanol would not be rate limiting during the conversion of glucose by Z. mobilis and (ii) indicated that ethanol can serve as an in vivo marker of cytoplasmic volume changes. This was verified by monitoring for the first time the changes of both cytoplasmic volume and extracytoplasmic and cytoplasmic concentrations of alpha and beta anomers of D-glucose in cell suspensions of a microorganism. These findings may open up new possibilities for kinetic studies of ethanol and sugar transport in Z. mobilis and other organisms.