No abstract

Solubilization of heavy bovine heart mitochondria with Triton X-100 leads to the selective extraction of F0F1ATP synthase monomer and dimer in a 2:1 ratio, as revealed by blue native gel electrophoresis (BN-PAGE). Second dimensional SDS-PAGE and immunoblotting with IF1 and F1 antibodies following BN-PAGE show that both aggregation states of the ATP synthase contain IF1. The monomer/dimer ratio does not change in extracts from mitochondria subjected to different energy conditions accompanied by IF1 binding modulation or from submitochondrial particles differing in IF1 content. In addition, the usual monomer/dimer ratio is observed even in submitochondrial particles deprived of IF1. Histochemical staining for ATPase activity demonstrates that the dimer is inactive, irrespective of its IF1 content. It is concluded that in the membrane of bovine heart mitochondria the ATP synthase dimer is a stable inactive structure, whose formation is not mediated by IF1 binding.

Gram-negative anaerobes belonging to the genera Fusobacterium, Prevotella and Porphyromonas were investigated for the presence of tetQ and ermF, which have been shown to be spread by conjugal elements. One hundred isolates from either sites of infection or various body sites in healthy subjects were studied. PCR was used to detect tetQ, and DNA-DNA hybridization studies on EcoRI chromosomal digests were undertaken to detect the presence of tetQ and ermF. Antibiotic sensitivity assays were performed on selected isolates to detect tetracycline, erythromycin and penicillin resistance. Twenty Fusobacterium isolates lacked tetQ, and were tetracycline sensitive. Twenty per cent of Prevotella spp. isolates both from clinical specimens and from healthy subjects were found to possess tetQ. Of 20 Porphyromonas isolates tested, one (Porphyromonas levii) from a case of bacterial vaginosis was shown to possess tetQ in the chromosome. The presence of tetQ was always associated with tetracycline resistance. Four isolates of Prevotella melaninogenica and one isolate of Prevotella were ermF-positive, although expression of erythromycin resistance was not consistently associated with detection of this gene. Antibiotic resistance phenotypes of Prevotella isolates were shown to be related to specific chromosomal restriction patterns by hybridization studies: tetracycline resistance and tetracycline/erythromycin resistance are conferred by Bacteroides tetracycline-resistant ERL elements, whereas the tetracycline/penicillin resistance phenotype could be due to spread of elements identified in Prevotella only. Tetracycline/erythromycin-resistant and tetracycline/erythromycin/penicillin-resistant P. melaninogenica isolates were found in this study. It appeared that the presence of tetQ and ermF in Bacteroides and Prevotella contributed to the persistence of antibiotic resistance isolates within the host and to potential spread to other organisms through conjugal elements.