The effect of variations in lipid composition of high-density lipoprotein on its interaction with receptors on human fibroblasts

To test whether the altered lipid composition of high-density lipoprotein (HDL) particles influences their ability to interact with the HDL receptor on cultured fibroblasts, HDL3 isolated from normal and diabetic donors with different degrees of hypertriglyceridemia was subjected to binding competition, cholesterol efflux, and net cholesterol transport assays. When HDL3 particles from different subjects were incubated with cholesterol-loaded fibroblasts, the initial rates of cholesterol efflux from cells to HDL3 particles appeared to be an exclusive function of the relative ability of HDL3 to interact with the HDL receptor. Variation in lipid composition of HDL3 particles did not appear to have any significant influence on either the receptor-binding or the efflux-promoting abilities of HDL3. When the movement of cholesterol between cells and HDL3 particles was allowed to approach equilibrium, the lipid composition of HDL3 became an important factor in determining the net amount of cholesterol removed from cells, with cholesterol-deficient triacylglycerol-rich HDL3 particles having the best capacity to promote net transport of cholesterol from cells. These results suggest that the ability of HDL to bind to its cell-surface receptor, rather than variations in the lipid composition of the HDL particle, is the major determinant of cholesterol efflux from cells to HDL particles. However, the lipid composition of HDL as well as its receptor-binding activity determine the net amount of cholesterol transported from cells over long-term incubation.     

Escherichia coli DNA gyrase: genetic analysis of gyrA and gyrB mutations responsible for thermosensitive enzyme activity.

Escherichia coli gyrA43 and gyrB203 alleles conferring temperature-sensitive (ts) growth encoded Gly751-->Asp and Pro171-->Ser substitutions in the DNA gyrase A and B subunits, respectively. A plasmid-borne gyrA43 allele was genetically dominant over a chromosomal quinolone-resistant gyrA gene at 30 degrees C but not at 42 degrees C. These results and others confirm the ts phenotype of the mutation, the first to be identified in the C-terminal DNA binding/complex stabilizing domain of gyrase A protein. By contrast, the Pro171-->Ser mutation is located near the ATP-binding site of gyrase B protein and could interfere with energy coupling during DNA supercoiling. These data are discussed in regard to recently described gyrA(ts) mutations that affect the control of chromosome segregation.     

Double-blind comparison of tolamolol,propranolol, practolol,and placebo in the treatment of angina pectoris.

Forty-two patients with angina pectoris have completed a randomized, double-blind trial comparing tolamolol 100 mg and 200 mg with propranolol 80 mg, practolol 100 mg, and placebo, all given three times a day. Tolamolol 200 mg thrice daily was found to be equivalent to propranolol 80 mg thrice daily in anti-anginal efficacy. Anginal attack rates and trinitrin consumption were significantly reduced by all active treatments as compared with the placebo but tolamolol and propranolol were the most effective. Tolamolol 200 mg thrice daily was most effective in reducing blood pressure, while propranolol was most effective in reducing the resting heart rate. All treatments except the placebo significantly increased the amount of exercise which could be performed before angina appeared (exercise work), while tolamolol 200 mg thrice daily significantly reduced Robinson''s index when compared with all other active agents. The degree of S-T segment depression induced by exercise was significantly lessened by both tolamolol and propranolol but not by practolol or placebo. There was no difference in patient preference between tolamolol and propranolol but tolamolol at both dose levels was preferred to practolol. Both tolamolol and propranolol are potent adrenergic beta-receptor antagonists and equal in anti-anginal efficacy but tolamolol has the advantage of being cardioselective. It is superior to practolol.     

Holliday junction resolvase in Schizosaccharomyces pombe has identical endonuclease activity to the CCE1 homologue YDC2.

A novel Holliday junction resolving activity has been identified in fractionated cell extracts of the fission yeast Schizosaccharomyces pombe . The enzyme catalyses endonucleolytic cleavage of Holliday junction-containing chi DNA and synthetic four-way DNA junctions. The activity cuts with high specificity a synthetic four-way junction containing a 12 bp core of homologous sequences but has no activity on another four-way junction (with a fixed crossover point), a three-way junction, linear duplex DNA or duplex DNA containing six mismatched nucleotides in the centre. The major cleavage sites map as single nicks in the vicinity of the crossover point, 3' of a thymidine residue. These data indicate that the activity has a strong DNA structure selectivity as well as a limited sequence preference; features similar to the Holliday junction resolving enzymes RuvC of Escherichia coli and the mitochondrial CCE1 (cruciform-cuttingenzyme 1) of Saccharomyces cerevisiae. A putative homologue of CCE1 in S.pombe (YDC2_SCHPO) has been identified through a search of the sequence database. The open reading frame of this gene has been cloned and the encoded protein, YDC2, expressed in E.coli . The purified recombinant YDC2 exhibits Holliday junction resolvase activity and is, therefore, a functional S.pombe homologue of CCE1. The resolvase YDC2 shows the same substrate specificity and produces identical cleavage sites as the activity obtained from S. pombe cells. Both YDC2 and the cellular activity cleave Holliday junctions in both orientations to give nicks that can be ligated in vitro. The partially purified Holliday junction resolving enzyme in fission yeast is biochemically indistinguishable from recombinant YDC2 and appears to be the same protein.     

DNA gyrase gyrA mutations in ciprofloxacin-resistant strains of Staphylococcus aureus: close similarity with quinolone resistance mutations in Escherichia coli.

The gyrA genes isolated from three ciprofloxacin-resistant clinical isolates of Staphylococcus aureus carried codon 84 (serine----leucine) and/or codon 85 (serine----proline) mutations that were absent in pretreatment susceptible strains. These substitutions occur in a region of the gyrase A protein wherein directly analogous mutations of serine 83----leucine and alanine 84----proline in Escherichia coli confer quinolone resistance. Thus, DNA gyrase A subunit mutations are implicated in resistance to ciprofloxacin in S. aureus.     

DNA cloning and organization of the Staphylococcus aureus gyrA and gyrB genes: close homology among gyrase proteins and implications for 4-quinolone action and resistance.

Staphylococcus aureus gyrA and gyrB genes, which encode the DNA gyrase A and B proteins, have been isolated and found to map contiguously. DNA sequence analysis revealed close homology between the S. aureus gyrase subunits and their counterparts in Bacillus subtilis and Escherichia coli, including several conserved amino acid residues whose substitution in E. coli confers resistance to 4-quinolones. These results are discussed in regard to quinolone resistance mechanisms in S. aureus.