Cloning of the cysB gene of Erwinia carotovora subsp. carotovora, and the identification of its product

Summary The suicide vector pJB4JI was used to generate a range of Tn5-induced mutants of Erwinia carotovora subsp. carotovora (Ecc). One mutant, HC500, was a cysteine auxotroph which had a non-pectolytic, non-cellulolytic, non-proteolytic phenotype when grown under sulphate-limitation. The cysteine lesion of HC500 was shown to be analogous to the cysB mutation of Escherichia coli. The Ecc-cysB ⁺ gene product was identified as a protein of M_r 36000.     

Immunoaffinity-purified opiate receptor specifically binds the delta-class opiate receptor ligand, cis-(+)-3-methylfentanylisothiocyanate, SUPERFIT

Using an antibody generated against the opiate receptor on NG108-15 cells, we recently purified the putative receptor from this hybrid cell line. We herein report that the purified receptor complex specifically binds tritiated cis-(+)-3-methylfentanylisothiocyanate (SUPERFIT), with the predominant binding associated with a 58 kDa polypeptide chain. Consistent with these findings is the in situ labeling of a 58 kDa protein with [3H]SUPERFIT on NG108-15 cells.     

Environmental factors influencing the vertical migration of planktonic rotifers in a hypereutrophic tarn

The diel vertical migration of planktonic rotifers in a small, hypereutrophic tarn was investigated on four occasions in 1983. When the tarn was isothermal the rotifers were distributed throughout the water column. After stratification, the rotifers were confined to the top 1–2 m of oxygenated water. On all four dates the rotifers were aggregated at specific depths in the water column. On some occasions, the pattern of aggregation changed as the animals performed distinct diurnal migrations. Keratella cochlearis, K. quadrata and Polyarthra vulgaris usually followed the reverse migrations of the phytoplankton. In contrast, the movements of Anuraeopsis fissa were less pronounced and were associated with variations in the depth of the oxycline.     

Comparative organization of nitrogen fixation-specific genes from Azotobacter vinelandii and Klebsiella pneumoniae: DNA sequence of the nifUSV genes.

In the facultative anaerobe Klebsiella pneumoniae 17 nitrogen fixation-specific genes (nif genes) have been identified. Homologs to 12 of these genes have now been isolated from the aerobic diazotroph Azotobacter vinelandii. Comparative studies have indicated that these diverse microorganisms share striking similarities in the genetic organization of their nif genes and in the primary structure of their individual nif gene products. In this study the complete nucleotide sequence of the nifUSV gene clusters from both K. pneumoniae and A. vinelandii were determined. These genes are identically organized on their respective genomes, and the individual genes and their products exhibit a high degree of interspecies sequence homology.     

Characterization of the chicken oocyte receptor for low and very low density lipoproteins

The chicken oocyte receptor for low and very low density lipoproteins has been identified and characterized. Receptor activity present in octyl-beta-D-glucoside extracts of oocyte membranes was measured by a solid phase filtration assay, and the receptor was visualized by ligand blotting. The protein had an apparent Mr of 95,000 in sodium dodecyl sulfate-polyacrylamide gels under nonreducing conditions and exhibited high affinity for apolipoprotein B-containing lipoproteins, but not for high density lipoproteins or lipoproteins in which lysine residues had been reductively methylated. Binding of lipoproteins was sensitive to EDTA, suramin, and treatment with Pronase. In these aspects, the avian oocyte system was analogous to the mammalian low density lipoprotein receptor in somatic cells. Furthermore, a structural relationship between the mammalian and avian receptors was revealed by immunoblotting: polyclonal antibodies directed against the purified bovine low density lipoprotein receptor reacted selectively with the 95-kDa chicken receptor present in crude oocyte membrane extracts.     

The use of biochemical markers, serotype and fimbriation in the detection of Escherichia coli clones

Biochemical reactions, O and K serotypes and presence of P-fimbriae were analysed in 116 Escherichia coli strains isolated in blood cultures from patients with bacteraemia and in 99 faecal strains isolated from healthy individuals. By using biochemical typing, the strains could be grouped into six main clusters with similarity index less than 0.8 (Gower, 1971) and altogether 16 subclusters with similarity index 0.82-0.89. The most discriminating tests between the clusters were fermentation of D-tagatose, saccharose, salicin and sorbose. No single biochemical property could differentiate bacteraemic isolates from faecal strains, although strains isolated from blood were significantly more often found in certain subclusters, whereas other subclusters contained mainly control strains. Bacteraemic strains possessed P-fimbriae more often, especially strains isolated from patients with E. coli in the urine concomitantly with bacteraemia. Equally, no single reaction could separate P-fimbriated from non-P-fimbriated strains. D-Tagatose was fermented more often by the P-fimbriated strains; on the other hand, melibiose and lactose fermentation tests were less often positive. Certain O serotypes (O1, O4, O6, O7, O18 and O25) were more common among bacteraemic isolates than controls. K serotypes such as K1, K5 and K52 were also more frequent among blood isolates. We conclude that a combination of biochemical tests, fimbriation and serotyping might be used to identify potentially pathogenic clusters of E. coli.     

Regional differences in quantities of histochemically detectable mucosubstances in nasal, paranasal, and nasopharyngeal epithelium of the bonnet monkey

Inhaled irritants induce secretory cell hyperplasia in nasal epithelium of animals. To characterize this response histochemically it is first important to know the histochemical character and distribution of epithelial mucosubstance in the normal nasal cavity. An automated image analyzing method was used to detect and quantitate acidic, neutral, and sulfated mucosubstances in the epithelium lining the nasal and paranasal airways of eight bonnet monkeys. Tissue sections 2 micron thick from defined regions of these airways were stained with either alcian blue/periodic acid-Schiff to demonstrate acid and neutral mucosubstances or high iron diamine to demonstrate sulfated mucins. Respiratory epithelium covering maxilloturbinates had the largest volume of stainable mucosubstance per unit surface area of basal lamina, whereas the maxillary sinus epithelium had the least. There was a general anteroposterior increase in the quantity of total epithelial mucosubstance along the septal and lateral walls of the nasal cavity, and there was more acidic than neutral mucosubstance in the posterior nasal airway than in the anterior. Epithelial mucosubstance in the maxillary sinus was predominantly neutral. Therefore, we conclude that there are substantial regional quantitative differences in stainable mucosubstances in the primate nasal epithelium which must be considered when examining nasal mucosa for irritant-induced changes in epithelial mucins.     

Cell matrix adhesion-related proteins VLA-1 and VLA-2: regulation of expression on T cells

The mitogens phytohemagglutinin (PHA) and concanavalin A inhibited the appearance of the very late activation antigen (VLA)-1, but did not inhibit VLA-2 expression on cultured activated T cells. In contrast to diminished VLA-1 expression, mitogen treatment caused increased cell surface expression of other activation antigens such as T10, HLA-DR, interleukin 2 (IL 2) receptor, and 4F2, and greater cell proliferation. Conversely, when T cells were not repetitively restimulated with mitogen, these less proliferative "postactivated" T cells had elevated VLA-1 expression. The diminished expression of VLA-1 caused by PHA was reversible since subsequent removal of mitogen was associated with increased VLA-1, paralleled by a decrease in interleukin 2 receptor levels. In addition to preventing or delaying the initial appearance of VLA-1, PHA stimulation also was somewhat effective in causing the disappearance of VLA-1 already present, especially on recently established cultures. However, cultures that had either never seen PHA, not seen PHA for several weeks, or been stimulated regularly with PHA, but were several months old, did not lose VLA-1 in response to PHA stimulation, suggesting that a state of insensitivity to PHA effects could be attained. Unlike PHA-stimulated T cells, T cells repetitively restimulated with alloantigen or the monoclonal antibody T3 did not show a marked absence of VLA-1 but rather showed an increased level of VLA-2 relative to VLA-1. Taken together, results of stimulation by either mitogen, alloantigen, or anti-T3 monoclonal antibody support the conclusion that T cell stimulation in general can cause a decreased VLA-1:VLA-2 ratio, whether by decreased VLA-1 or increased VLA-2. These shifts in VLA-1:VLA-2 ratios are probably not simply the result of shifts in the relative proportions of different subpopulations, because similar growth-related changes in this ratio were observed on the T cell line ANITA, which is a homogeneous population of cells. Because both VLA-1 and VLA-2 are differentially regulated on cultured, long term activated T cells depending on stage of activation and growth conditions, and are members of a family of at least five heterodimers that includes cell matrix adhesion molecules, we suggest that these studies will provide clues to novel aspects of T cell growth regulation, perhaps relating to T cell-matrix adhesion.     

Mitochondrial DNA of the extinct quagga: Relatedness and extent of postmortem change

Sequences are reported for portions of two mitochondrial genes from a domestic horse and a plains zebra and compared to those published for a quagga and a mountain zebra. The extinct quagga and plains zebra sequences are identical at all silent sites, whereas the horse sequence differs from both of them by 11 silent substitutions. Postmortem changes in quagga DNA may account for the two coding substitutions between the quagga and plains zebra sequences. The hypothesis that the closest relative of the quagga is the domestic horse receives no support from these data. From the extent of sequence divergence between horse and zebra mitochondrial DNAs (mtDNAs), as well as from information about the fossil record, we estimate that the mean rate of mtDNA divergence in Equus is similar to that in other mammals, i.e., roughly 2% per million years.     

d-Aspartate in Human Brain

The presence of the biologically uncommon D-aspartic acid (D-aspartate) in human brain white matter has been previously reported. The earlier study has now been expanded to include D/L-aspartate ratios from 67 normal brains. The data show that the D-aspartate content increases rapidly from 1 year to approximately 35 years of age, levels off in middle age, and then appears to decrease somewhat. The D-aspartate content in gray matter remains at a consistently low level (half of that found in white matter) throughout the human life span. Within the limitations of current analytical methods, there was no detectable difference in D/L-aspartate ratios in white and gray matter of brains with Alzheimer's disease and several other pathologies when compared with brains of normal subjects. However, the presence of a significant D-aspartate level in white matter during the adult life span may lead to changes in protein configuration related to dysfunctions associated with the aging brain.