The interaction in vivo of transferrin and asialotransferrin with liver cells.

Rat transferrin or asialotransferrin doubly radiolabelled with 59Fe and 125I was injected into rats. A determination of extrahepatic and hepatic uptake indicated that asialotransferrin delivers a higher fraction of the injected 59Fe to the liver than does transferrin. In order to determine in vivo the intrahepatic recognition sites for transferrin and asialotransferrin, the liver was subfractionated into parenchymal, endothelial and Kupffer cells by a low-temperature cell isolation procedure. High-affinity recognition of transferrin (competed for by an excess of unlabelled transferrin) is exerted by parenchymal cells as well as endothelial and Kupffer cells with a 10-fold higher association (expressed per mg of cell protein) to the latter cell types. In all three cell types iron delivery occurs, as concluded from the increase in cellular 59Fe/125I ratio at prolonged circulation times of transferrin. It can be calculated that parenchymal cells are responsible for 50-60% of the interaction of transferrin with the liver, 20-30% is associated with endothelial cells and about 20% with Kupffer cells. For asialotransferrin a higher fraction of the injected dose becomes associated with parenchymal cells as well as with endothelial and Kupffer cells. Competition experiments in vivo with various sugars indicated that the increased interaction of asialotransferrin with parenchymal cells is specifically inhibited by N-acetylgalactosamine whereas mannan specifically inhibits the increased interaction of asialotransferrin with endothelial and Kupffer cells. Recognition of asialotransferrin by galactose receptors from parenchymal cells or mannose receptors from endothelial and Kupffer cells is coupled to active 59Fe delivery to the cells. It is concluded that, as well as parenchymal cells, liver endothelial and Kupffer cells are also quantitatively important intrahepatic sites for transferrin and asialotransferrin metabolism, an interaction exerted by multiple recognition sites on the various cell types.     

Die Entwicklung der Laut?u?erungen bei der Graugans (Anser anser)

Zusammenfassung Das Lautrepertoire von sechs handaufgezogenen Graugänsen wurde in individuellen Längsschnitten vom Schlüpfen bis zum Alter von 100 Tagen mit sonagraphischer Methode analysiert. Die Einteilung des Repertoires erfolgte nach drei verschiedenen Kriterien: Klang, Sonagramm und Kontext. Im jugendlichen Repertoire ließen sich 4 Rufklassen voneinander trennen. Im Laufe der Jugendentwicklung veränderten sich vor allem die phonetischen Parameter der Rufe, während die syntaktischen weitgehend konstant blieben. Nach einem Anstieg in der ersten Woche fiel die Tonhöhe der Rufe bei allen untersuchten Individuen und Rufklassen gleichmäßig ab. Der Stimmbruch, meßbar an der Zunahme geräuschhafter Komponenten, war ein kontinuierlicher, langfristiger Prozeß. Die Individualität war nicht in einem einzelnen, wohl aber in der Kombination mehrerer Merkmale faßbar. Die Geschlechter waren schon früh an Unterschieden in der Tonhöhe erkennbar. An sonstigen ontogenetischen Veränderungen innerhalb des Untersuchungszeitraumes konnten die Erweiterung des Repertories durch Aufspaltung und Reifung sowie ein möglicher Verlust von Rufklassen beobachtet werden.

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Juvenile development of vocalizations in Greylag Geese (Anser anser)

Summary The juvenile vocal development of six Greylag Geese from hatching up to the age of 100 days was analysed by using sonagraphic methods. The calls were classified by means of the acoustic phenomenon, sonagrams, and context criteria. The juvenile repertoire included four main call types. During ontogeny, changes of call parameters mainly referred to phonetic characteristics, whereas syntactic parameters remained more or less constant. The frequency pitch of calls changed to higher levels during the first week of development while in older goslings this parameter decreased constantly. This held true for each individual and call type. Breaking of the voice, defined as an increment of noisy parts in the vocalizations, is described as a continuous long term process. Structural individuality of calls apparently was not based on one single parameter, but on the combination of several ones. Sex differences were found in the pitch of vocalizations, correlating with different body weights of male and female geese. Additional developmental processes of the vocal repertoire included differentiation, maturation of new call types, and apparent suppression of juvenile ones. The functional organization of the juvenile call system in Greylags is discussed.

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Herrn Dr. Hans Löhrl zum 75. Geburtstag gewidmet     

Purification, properties, and N-terminal amino acid sequence of homogeneous Escherichia coli 2-amino-3-ketobutyrate CoA ligase, a pyridoxal phosphate-dependent enzyme

Starting with 100 g (wet weight) of a mutant of Escherichia coli K-12 forced to grow on L-threonine as sole carbon source, we developed a 6-step procedure that provides 30-40 mg of homogeneous 2-amino-3-ketobutyrate CoA ligase (also called aminoacetone synthetase or synthase). This ligase, which catalyzes the cleavage/condensation reaction between 2-amino-3-ketobutyrate (the presumed product of the L-threonine dehydrogenase-catalyzed reaction) and glycine + acetyl-CoA, has an apparent molecular weight approximately equal to 85,000 and consists of two identical (or nearly identical) subunits with Mr = 42,000. Computer analysis of amino acid composition data, which gives the best fit nearest integer ratio for each residue, indicates a total of 387 amino acids/subunit with a calculated Mr = 42,093. Stepwise Edman degradation provided the N-terminal sequence of the first 21 amino acids. It is a pyridoxal phosphate-dependent enzyme since (a) several carbonyl reagents caused greater than 90% loss of activity, (b) dialysis against buffer containing hydroxylamine resulted in 89% loss of activity coincident with an 86% decrease in absorptivity at 428 nm, (c) incubation of the apoenzyme with 20 microM pyridoxal phosphate showed a parallel recovery (greater than 90%) of activity and 428-nm absorptivity, and (d) reduction of the holoenzyme with NaBH4 resulted in complete inactivation, disappearance of a new absorption maximum at 333 nm. Strict specificity for glycine is shown but acetyl-CoA (100%), n-propionyl-CoA (127%), or n-butyryl-CoA (16%) is utilized in the condensation reaction. Apparent Km values for acetyl-CoA, n-propionyl-CoA, and glycine are 59 microM, 80 microM, and 12 mM, respectively; the pH optimum = 7.5. Added divalent metal ions or sulfhydryl compounds inhibited catalysis of the condensation reaction.     

Comparison of the three-dimensional structure of two human rhinoviruses (HRV2 and HRV14)

An attempt has been made to build a model of human rhinovirus 2 (HRV2) based on the known human rhinovirus 14 (HRV14) structure. HRV2 was selected because its amino acid sequence is known and because it belongs to the minor rhinovirus receptor class as compared to HRV14, which belongs to the major class. Initial alignment of HRV2 with HRV14 based on the primary sequence and the knowledge of the three-dimensional structure of HRV14 showed that the most probable position of the majority of insertions and deletions occurred in the vicinity of the neutralizing immunogenic sites (NIm). Out of a total of 855 amino acids present in one copy of each of the capsid proteins VP1 through VP4 of HRV14, 411 are different between the two viruses. There are also 6 amino acid residues inserted and 14 residues deleted in HRV2 relative to HRV14. Examination of amino acid interactions showed several cases of conservation of function, e.g., salt bridges or the filling of restricted space. The largest variation amongst the residues lining the canyon, the putative receptor binding site, was in the carboxy-terminal residues of VP1.     

Detection by ELISA of Two Tobamoviruses in Orchids Using Monoclonal Antibodies

Five monoclonal antibodies (McAbs) were raised to the tobamovirus, odontoglossum ringspot virus (ORSV). All five McAbs reacted with the virus in double antibody sandwich (DAS) ELISA but not in an ELISA using virus-coated plates. All the McAbs recognized a panel of ORSV strains and isolates, although one of the antibodies reacted better with some isolates and another reacted less with certain isolates than with type ORSV. It was possible to use the same McAbs both as coating and as biotinylated antibody in DAS-ELISA. None of the five McAbs was able to bind to orchid strains of tobacco mosaic virus (TMV). In order to detect strains of both viruses, ORSV and TMV, in infected orchids it was necessary to include also McAbs raised against TMV in the immunoassays. The use of a mixed polyclonal-monoclonal antibody DAS-ELISA system is advocated for detecting both tobamoviruses in orchids.     

Mono- through hexanucleotide composition of the Escherichia coli genome: a Markov chain analysis.

Several statistical methods were tested for accuracy in predicting observed frequencies of di- through hexanucleotides in 74,444 bp of E. coli DNA. A Markov chain was most accurate overall, whereas other methods, including a random model based on mononucleotide frequencies, were very inaccurate. When ranked highest to lowest abundance, the observed frequencies of oligonucleotides up to six bases in length in E. coli DNA were highly asymmetric. All ordered abundance plots had a wide linear range containing the majority of the oligomers which deviated sharply at the high and low ends of the curves. In general, values predicted by a Markov chain closely followed the overall shape of the ordered abundance curves. A simple equation was derived by which the frequency of any nucleotide longer than four bases in the E. coli genome (or any genome) can be relatively accurately estimated from the nested set of component tri- and tetranucleotides by serial application of a 3rd order Markov chain. The equation yielded a mean ratio of 1.03 +/- 0.94 for the observed-to-expected frequencies of the 4,096 hexanucleotides. Hence, the method is a relatively accurate but not perfect predictor of the length in nucleotides between hexanucleotide sites. Higher accuracy can be achieved using a 4th order Markov chain and larger data sets. The high asymmetry in oligonucleotide abundance means that in the E. coli genome of 4.2 X 10(6) bp many relatively short sequences of 7-9 bp are very rare or absent.     

Vascular plants first described in Rydberg’s flora of colorado

Types are indicated for 28 new names inadvertently published in Rydberg’sFlora of Colorado. Included are a discussion as to the validity of these new names and an explanation of the causes of this oversight. A lectotype ofLupinus decumbens var.argentatus is herein designated by Rupert C. Barneby.     

Bactericidal effect of lactoferrin on Legionella pneumophila: effect of the physiological state of the organism

Lactoferrin has been previously shown to be bactericidal for Legionella pneumophila. The current study showed that CaCl2, Mg(NO3)2, and MgCl2, but not NaCl, blocked killing. Activity was pH dependent with the greatest activity at 5.0. Sensitivity of the organism was dramatically affected by the growth conditions. Log phase 12 h, broth-grown cells were most sensitive, with older cultures becoming more resistant. Plate-grown cells were completely resistant. Lactoferrin binding, as detected by immunofluorescence microscopy, was temperature dependent (no binding at 4 degrees C), but was independent of killing.     

Secondary transport of amino acids by membrane vesicles derived from lactic acid bacteria

Lactococci are fastidious bacteria which require an external source of amino acids and many other nutrients. These compounds have to pass the membrane. However, detailed analysis of transport processes in membrane vesicles has been hampered by the lack of a suitable protonmotive force (pmf)-generating system in these model systems. A membrane-fusion procedure has been developed by which pmf-generating systems can be functionally incorporated into the bacterial membrane. This improved model system has been used to analyze the properties of amino acid transport systems in lactococci. Detailed studies have been made of the specificity and kinetics of amino acid transport and also of the interaction of the transport systems with their lipid environment. The properties of a pmf-independent, arginine-catabolism specific transport system in lactococci will be discussed.Abbreviations pmf protonmotive force - transmembrane electrical potential - pH transmembrane pH gradient - PE phosphatidylethanolamine - PC phosphatidylcholine Paper adapted from a treatise Secondary Transport of Amino Acids by Membrane Vesicles Derived from Lactic Acid Bacteria and awarded the Kluyver Prize 1988 by the Netherlands Society of Microbiology.