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J Smit

Publications and source records attributed to J Smit.

102 records · Page 6Linked to original sources
PubMed1984

Cloning of the major protein of the Caulobacter crescentus periodic surface layer: detection and characterization of the cloned peptide by protein expression assays.

A precisely ordered crystalline array is found on the surface of the bacterium Caulobacter crescentus CB15. Using an immunological assay, we identified recombinant bacteriophage clones expressing the predominant protein of this structure from a lambda 1059 library of C. crescentus CB15 DNA. A single 4.4-kilobase HindIII fragment encoded a polypeptide whose antigenic determinants, molecular weight, and peculiar solubilization properties were identical with those of the authentic predominant polypeptide (130K) of the surface array. The 130K protein was produced as a discrete product as a result of gene transcription initiated from a lambda promoter; several experiments suggested that the Caulobacter promoter for this gene is not efficiently recognized by the Escherichia coli transcription machinery. Genomic Southern analysis revealed a single copy of the 130K protein gene per genome. The 130K protein gene was hybridized with DNA of two closely related laboratory strains of C. crescentus which have lost their ability to produce a surface array. One of these strains, CB2, possesses an homologous copy of the 130K gene, whereas DNA from the other strain, CB13B1a, showed a lesser degree of hybridization to the 130K gene probe; genomic fragments which did hybridize were of different sizes in CB13 as compared with those of CB15. These findings are discussed in relation to studies of the surface array function and its role in cellular morphogenesis in this stalk-forming bacterium.

Bacterial Proteins↗
PubMed1982

Cell surface patterning and morphogenesis: biogenesis of a periodic surface array during Caulobacter development.

Shape changes, extended processes, and other surface elaborations are associated with cellular differentiation, and the cell membranes involved with these developmental changes often are reshaped without a major alteration in biochemical composition. Caulobacter crescentus produces a hexagonally-packed periodic surface layer that covers the entire cell and further, mimics some of the membrane-mediated changes of higher organisms by forming a membranous stalk during its distinctive life cycle. Growth of the surface layer was examined during the cell cycle by treating synchronously growing cells with surface layer antibody, continuing growth, and then labeling for electron microscopy with a protein A-colloidal gold conjugate. Three regions of distinctive surface array biogenesis were resolved. The periodic surface layer on the main cell body was enlarged by insertion of new material at numerous uniformly distributed points. In contrast, the surface layer on the stalk appeared as entirely new synthesis. In examining growth of the stalk in subsequent generations, we noted that growth of stalk surface persisted at the stalk-cell body junction. The region of cell division also showed a pattern of entirely new surface layer production at late stages in division, similar to the stalk. The immunocytological method also facilitated a careful examination of stalk initiation and growth. Although initiation was under precise temporal and spatial regulation, the rate of stalk elongation was variable from cell to cell and apparently no longer under cell cycle control. The similarity of surface layer biogenesis on the stalk and the site of cell division may be a significant reflection of other events occurring at the cell pole. A model suggested by this and other studies that can account for the temporal pattern of polar morphogenesis is discussed, as is the potential relationship between the geometrically ordered surface array and the formation or maintenance of the stalk.

Antigens, Bacterial↗
PubMed1982

Indoor air pollution by carbon monoxide and nitrogen oxides.

The results of an exploratory survey of indoor levels of carbon monoxide and nitrogen dioxide from gas-fired cooking and waterheating appliances in the Dutch cities of Arnhem and Enschede in the fall of 1980 are reported. Measurements were carried out electrochemically (Ecolyzer 2000) or with Draeger tubes in the case of CO and with Palmes diffusion tubes (5 to 8 days exposure) in the case of NO2. For CO, in 18% (27%) of the homes visited the limit of 600 (300) ppm in the flue gases was exceeded, whereas the Dutch Installation Code Standard of 50 ppm CO in room air was exceeded in 17% of the homes. The arithmetic mean value of the NO2 concentration in 286 homes was 118 micrograms/m3 with a range of 35 to 472 micrograms/m3. The corresponding figures for living rooms were 58 and 35 to 346 micrograms/m3, respectively. Outdoor NO2 concentrations were 2 to 3 times lower than indoor concentrations.

Air Pollutants↗
PubMed1981

Caulobacter crescentus pilin. Purification, chemical characterization, and NH2-terminal amino acid sequence of a structural protein regulated during development.

The pili of Caulobacter crescentus are assembled during swarmer cell development at the differentiating cell pole. Under specific growth conditions, it was found that C. crescentus CB15 will produce insoluble pigmented granules which entrap pili lost during cell growth along with other extracellular proteins. This provided a strategy for protein purification. Pilin was purified from these granules by gel filtration in the presence of high levels of detergent. A protein of the appropriate molecular weight for pilin was isolated by this procedure and demonstrated to be pilin by specific labeling of the intact pilus with ferritin-coupled antibodies. CB15 pilin has an apparent molecular weight of 8,000, is rich in hydrophobic amino acids (greater than 70%), and has a broad isoelectric focusing range centered about a pI of 6.6. Periodic acid-Schiff reagent staining did not demonstrate carbohydrate modification of the monomer. The pilins of both CB15 and a related strain CB13 cross-react with anti-CB15 pilin antibody. Both strains also demonstrate similar RNA bacteriophage sensitivity, although there were significant differences in the amino acid composition, molecular weight, and other physical properties of the pilins from these two related strains. The NH2-terminal amino acid sequence of about 40% of the CB15 pilin molecular has been determined and bears some resemblance to that of some common pili; however, the sequences are not homologous and there is no indications of an unusual NH2-terminal amino acid in Caulobacter pilin.

Amino Acid Sequence↗
PubMed1981

Periodic surface array in Caulobacter crescentus: fine structure and chemical analysis.

A periodic array structure on the cell surface of Caulobacter crescentus CB15 was revealed by electron microscopy of the cell envelope, using negative staining, thin-sectioning, and freeze-etching. This structural layer has been isolated from liquid cultures, in which large pieces of the two-dimensional array are shed by cells grown to high density. Often areas of intact array corresponding to the entire cell surface could be found. The hexagonally arranged structure was highly ordered and had an unusual degree of complexity, as determined by optical diffraction and computer processing of micrographs of negatively stained, isolated surface array. Filtered, reconstructed images were obtained from both normal and low-electron-dose micrographs demonstrating resolutions of 2.9 and 25 nm, respectively. Comparison by optical diffraction and image filtering of micrographs recorded by using either normal or minimal beam exposure techniques suggested that the lower-resolution features of the image are very stable to electron exposure. Gel electrophoresis indicated that isolated array preparations contain a number of polypeptides. It appears likely that more than one of these proteins are structural components of the array, in contrast to a single protein found in many bacterial surface arrays. The Caulobacter surface array is also unusual in that the repeated units are widely spaced with no apparent direct connection. Computer spatial averaging provided information about the shape and complexity of the connecting elements, and this was compared with some additional electron microscopic evidence of linking structures. Thin-sectioning studies confirmed the image features seen by other techniques, but the addition of tannic acid in the fixation procedure was required to visualize the structure. A comparison of these results with out current knowledge of the Caulobacter cell envelope suggests interesting questions about the biogenesis of this membrane structure and its involvement in the cell development process of this organism.

Bacteria↗
PubMed1978

Outer membrane of gram-negative bacteria. XVIII. Electron microscopic studies on porin insertion sites and growth of cell surface of Salmonella typhimurium.

Salmonella typhimurium contains three "major proteins" or "porins" (34K, 35K, and 36K) in the outer membrane. A mutant strain producing only the 35K porin was first grown in media containing high concentrations of NaCl to "repress" the porin synthesis and then was shifted into a medium without NaCl. The newly made porin molecules were then labeled with the ferritin-coupled antibody at various times after the shift, and the samples were examined by whole-mount, freeze-etching, and thin-section electron microscopy. These experiments showed that newly inserted porins appeared as discrete patches uniformly distributed over the surface of the cell and, furthermore, that the sites of adhesion between the inner and outer membrane were most probably the pathway by which the newly made porin molecules appeared on cell surface. The 34K and 36K porins were also inserted in the same manner, since the appearance of new porins at discrete sites all over the cell surface was also observed when cells with wild-type porin phenotype were treated with unlabeled antibody to block existing antigenic sites, subsequently regrown, and labeled with the ferritin-coupled antibody. Since porins comprise a major portion of the densely packed, relatively immobile, "protein framework" of the outer membrane, these results lead us to conclude that the outer membrane grows predominantly by diffuse intercalation rather than by the zonal growth mechanism.

Antibodies, Bacterial↗
PubMed1978

Pneumothorax in the Ehlers-Danlos syndrome: consequence of coincidence?

A 25 year-old man, known to suffer from Ehlers-Danlos syndrome, developed a left-sided pneumothorax. At thoracotomy, pulmonary blebs and bullae were found at the apex of the lung. Bullectomy with removal of adjacent lung tissue was performed. The histological and ultrastructural findings in the lung tissue must be regarded as non-specific. These findings do not reflect a direct cause-and-effect relationship, even in the presence of the inheritable connective tissue disorder.

Adult↗
PubMed1975

Outer membrane of Salmonella typhimurium: chemical analysis and freeze-fracture studies with lipopolysaccharide mutants.

The outer membrane layer of the cell wall was isolated from wild-type Salmonella typhimurium LT2 as well as from its mutants producing lipopolysaccharides with shorter saccharide chains. Chemical analysis of these preparations indicated the following. (i) The number of lipopolysaccharide molecules per unit area was constant, regardless of the length of the saccharide side chain in lipopolysaccharide. (ii) In contrast, in "deep rough" (Rd or Re) mutants producing the lipopolysaccharides with very short saccharide chains, the amount of outer membrane protein per unit surface area decreased to about 60% of the value in the wild type. (iii) In the wild type, the amount of phospholipids is slightly less than what is needed to cover one side of the membrane as a monolayer. In comparison with the wild type, the outer membrane of Rd and Re mutants contains about 70% more phospholipids, which therefore must be distributed in both the outer and inner leaflets of the membrane. Freeze-fracture studies showed that the outer membrane of Re mutants were easily fractured, but fracture became increasingly difficult in strains producing lipopolysaccharides with longer side chains. The convex fracture face was always nearly smooth, but the concave fracture face or the outer half of the membrane was densely covered with particles 8 to 10 nm in diameter. The density of particles was decreased in Re mutants to the same extent as the reduction in proteins, suggesting the largely proteinaceous nature of particles. A model for the supramolecular structure of the outer membrane is presented on the basis of these and other results.

Bacterial Proteins↗
PubMedDate not supplied

Divided attention years after severe closed head injury: the effect of dependencies between the subtasks.

Lesions of white matter which connects distant brain areas are characteristic for closed head injury (CHI). It was predicted that this impairs divided attention only if dependent subtasks are used which require communication between corresponding brain processes. Fourteen chronic severe CHI patients (mean age 42 years) and 14 healthy controls, matched on age, gender, and education, participated in an experiment on divided attention. A typical left hemisphere task and a typical right hemisphere task must be simultaneously performed in a condition where they are completely independent and in conditions with dependencies between the tasks. Only in the dependent conditions the CHI group showed poor divided attention performance.

Adult↗