Studies in vitro of the nature and synthesis of the cell wall ofChlamydia trachomatis

The possibility thatChlamydia trachomatis contains peptidoglycan was examined by three methods. Preincubation of chlamydia with enzymes known to cleave peptidoglycan had no adverse effect on the subsequent development. Immunofluorescence studies with antistreptococcal peptidoglycan antisera failed to show any cross reactions with chlamydial antigens. The antichlamydial activity of anti-cell-wall antimicrobials was examined; lactams proved the most active, and cycloserine and bacitracin also showed antichlamydial activity. Alaphosphin, phosphomycin, and vancomycin showed no antichlamydial activity at the concentrations tested.     

A radioimmunoassay for 6-keto- prostaglandin F1α

A radioimmunoassay for 6-keto-prostaglandin F_1α has been developed. The assay is accurate and sensitive but since the antiserum cross-reacts 5–10% with prostaglandins (PGs) of the E and F series, solvent extraction and thin layer chromatography are required fo absolute specifity. The assay has been validated by comparison with a radiochemical assay and by the use of an inhibitor of 6-keto PGF_1α formation, 15-hydroperoxy arachidonic acid. 6-Keto PGF_1α was found to have a low cross reaction with antisera directed against PGE₂, PGF_2α and thromboxane B₂.     

Identification of polypeptides necessary for chemotaxis in Escherichia coli.

Molecular cloning techniques were used to construct Escherichia coli-lambda hybrids that contained many of the genes necessary for flagellar rotation and chemotaxis. The properties of specific hybrids that carried the classical "cheA" and "cheB" loci were examined by genetic complementation and by measuring the capacity of the hybrids to direct the synthesis of specific polypeptides. The results of these tests with lambda hybrids and with a series of deletion mutations derived from the hybrids redefined the "cheA" and "cheB" regions. Six genes were resolved: cheA, cheW, cheX, cheB, cheY, and cheZ. They directed the synthesis of specific polypeptides with the following apparent molecular weights: cheA, 76,000 and 66,000; cheW, 12,000; cheX, 28,000; cheB, 38,000; cheY, 8,000; and cheZ, 24,000. The presence of another gene, cheM, was inferred from the protein synthesis experiments. The cheM gene directed the synthesis of polypeptides with apparent molecular weights of 63,000, 61,000, and 60,000. The synthesis of all of these polypeptides is regulated by the same mechanisms that regulate the synthesis of flagellar-related structural components.     

Macromolecular Composition of Spores from the Filamentous Cyanobacterium Anabaena cylindrica

Spores were isolated from the filamentous cyanobacterium Anabaena cylindrica, and their deoxyribonucleic acid, ribonucleic acid, and protein compositions were determined.     

Phase variation: genetic analysis of switching mutants

Site-specific inversion of a controlling element is responsible for flagellar phase transition in Salmonella. When a 900 bp DNA sequence is in one configuration, it allows the expression of the H2 gene, a structural gene which codes for the flagellar antigen. When it is in the opposite configuration, the H2 gene is not expressed. A hybrid λ phage containing the invertible control region and the adjacent H2 gene was constructed, and expression of the H2 gene was shown to be regulated by the orientation of the inversion region. Transposon Tn5 insertion derivatives of this hybrid phage were isolated and λH2::Tn5 mutants defective for inversion (H2 switching) were selected and characterized. Two classes of switching phenotypes were observed among the mutants—those which had slightly reduced frequencies of transition from expression of the H2 gene (H2 on) to nonexpression (H2 off) (intermediate class) and those in which the frequency of transition was reduced at least three orders of magnitude (null class). Physical mapping of the Tn5 insertion sites revealed that in all mutants the insertion was located inside the inversion region. Tn5 insertion sites in the null class of mutants defined a region of DNA including approximately 500 bp which was necessary for inversion. Genetic complementation tests showed that these λH2::Tn5 mutants could invert the H2 gene control element if the 500 bp region was introduced in the trans configuration. It is concluded that a gene is located inside the inversion segment and codes for a protein which is required for the inversion event. Furthermore, the two sites at which the crossover event occurred functioned in a cis configuration and were required for inversion. The presence of a gene which is involved in controlling site-specific recombination events may be a general feature of transposon-like elements.     

Solubility of carbon dioxide in lipid bilayer membranes and organic solvents

Partition coefficients of carbon dioxide into lipid bilayers (liposomes) and organic solvents were measured as a function of temperature. The molar partition coefficient of CO2 into liposomes of egg lecithin at 25 degrees C was 0.95 (ml CO2/ml lipid)/(ml CO2/ml saline). The addition of an equimolar amount of cholesterol to the egg lecithin decreased the partition coefficient by about 25%. The partition coefficients for CO2 into liposomes at 25 degrees C were lower than the partition coefficients into octanol (1.3), hexadecane (1.5) and olive oil (1.7). The results are discussed in terms of the solubility-diffusion model of non-electrolyte transport through lipid bilayer membranes.     

Use of RP4-prime plasmids constructed in vitro to promote a polarized transfer of the chromosome in Escherichia coli and Rhizobium meliloti.

Summary RP4-prime plasmids containing chromosomal fragments of either Escherichia coli or Rhizobium meliloti were constructed in vitro. When introduced into E. coli or R. meliloti respectively, they promoted a polarized transfer of the chromosome as demonstrated either by the gradient of transfer of various markers or by the study of the genetic constitution of recombinants. In E. coli, mobilization was shown to be dependent upon the presence of a functional rec A system. Inheritance of markers was due to their integration into the chromosome of the recipient as shown by the need for a functional rec A system in the recipient E. coli or by mobilization of recessive markers in R. meliloti. The system described could be applied to genetic mapping in any Gram negative bacteria.     

The biliary excretion of trypsin in rats

The serum half-life of bovine [³H]acetyltrypsin was estimated to be 9 rain following intravenous administration in rats. This was maintained when six successive doses of 200 g each were given at 1-h intervals. The enzyme was removed from the circulation after complexing with 2-macroglobulin (2-M). The amount of³H label appearing in bile increased with each successive dose and this was associated with breakdown products (<10 000 daltons) of the ₂–M/[³H] acetyltrypsin. Intact –M/[³H] acetyltrypsin was recovered from bile but represented only 0.06% of the administered dose of active enzyme.