Inhibition of cation cotransport by cholesterol enrichment of human red cell membranes.

1. Human red cells were enriched with cholesterol by incubation with lipid dispersions having a high cholesterol: phospholipid mol ratio and the kinetics of the furosemide-sensitive cotransport for Na+ and K+ were measured. 2. Influxes of both K+ and Na+ through this system were inhibited by 70 and 76% in cholesterol-rich cells (cholesterol: phospholipid mol ratio 1.80) and the Km of the furosemide-sensitive flux components for both K+ and Na+ decreased. 3. Effluxes of both K+ and Na+ are inhibited by furosemide and the magnitudes of these furosemide-sensitive components are markedly decreased in cholesterol-rich cells. 4. The inhibitory effect of cholesterol enrichment on this carrier-mediated transport of cations suggests that cholesterol may either alter the position of the carrier or retard its movement within a more viscous membrane micro-environment.     

Effects of subunit V antibodies on the topology of the subunit and the activity of beef heart cytochrome-c oxidase

Redox-sensitive epitopes on subunit V of beef heart cytochrome-c oxidase were demonstrated previously using polyclonal subunit-specific antibodies raised in rabbits. The antibodies only slightly inhibited electron transfer, and the accessibility of their epitopes depended on the presence of a membrane and on the redox state of the oxidase. The present paper describes additional preparations of antibodies raised against subunit V. These antibodies have an even higher subunit specificity, they are more than three times as inhibitory against electron transfer, and their binding does not require a membrane. Moreover, the redox-sensitive nature of their binding to detergent-dispersed oxidase is sensitive to the method of its isolation. We discuss inferences that can be drawn from a detailed quantitative comparison of the interactions of the two antibody preparations with the antigen in different environments. The techniques used in the comparison can be used to examine other perturbants of the oxidase as to their effects on specific segments of the enzyme.     

A critical analysis of excretory products as sex controlling agents in cladocera

Summary Conditions seem to exist under which the sex of the young to be produced by a Cladoceran mother is not determined by the animal's immediate environment. In such mothers a sex determining mechanism appears to have been inherited, and such mothers have been called Sex Fast.Laboratory stock cultures of Cladocera not infrequently pass through periods of depression. While in such periods the mothers are for the most part Sex Fast. Whether this can be termed sex control by inherent sexual cycles is questionable.The contention that many factors in the external environment of Cladocera may under suitable conditions act as sex controlling factors has been conclusively proven by numerous investigators. For the most part these are factors which affect the mother's metabolism, presumably accelerating or retarding the mother's development, acceleration being associated with female production, retardation with male production.Food, temperature and excretory substances are the external environmental factors most often associated with sex control in Cladocera. The evidence supporting the rôle of food and temperature in this reaction appears to be adequate but further experimental evidence supporting excretory substances in the control of sex is needed before a specific rôle in sex control in Cladocera can be assigned to this factor.

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Zusammenfassung Wahrscheinlich gibt es Umstände, unter denen das Geschlecht der Nachkommenschaft einer Cladocera von der unmittelbaren Umgebung unabhängig ist. Bei solchen Muttertieren scheint ein geschlechtsbestimmender Mechanismus geerbt zu sein, und daher hat man solche Mütter sexfast genannt.Die gewöhnlichen Laboratorienkulturen der Cladocera machen häufig Perioden der Erschöpfung durch. Während solcher Perioden sind die Mütter meistens sexfast. Ob man diese Erscheinung als Geschlechtsbestimmung durch vererbten Geschlechtszyklus bezeichnen darf, ist fraglich.Die Behauptung, daß viele Faktoren in der Umgebung der Cladocera unter günstigen Verhältnissen als geschlechtsbestimmend wirksam sind, ist schon von vielen Forschern bewiesen worden. Diese Faktoren sind meistens solche, die den Stoffwechsel der Mutter beeinflussen. Wahrscheinlich üben sie einen beschleunigenden oder verzögernden Einfluß auf die Entwicklung der Mütter aus. Im ersteren Falle glaubt man eine größere Anzahl weiblicher, im letzteren männlicher Nachkommenschaft feststellen zu können.Ernährung, Stoffwechselprodukte und Temperatur sind die äußeren Faktoren, die man am häufigsten mit der Geschlechtsbestimmung bei der Cladocera in Zusammenhang bringt. Der Beweis für die Rolle von Ernährung und Temperatur bei dieser Reaktion scheint genügend, aber wir müssen einen weiteren experimentellen Beweis für die Rolle der Stoffwechselprodukte haben, ehe wir diesem Faktor eine spezifische Rolle in der Geschlechtsbestimmung bei Cladocera zuschreiben dürfen.

     

Antibody-independent C1 activation by E. coli

Antibody-independent interactions of C1 with several E. coli strains were examined. Purified C1 was directly activated by the semi-rough mutant E. coli J-5, its parental wild-type strain, E. coli 0111:B4, and two clinical isolates, E. coli (P) and E. coli (A), in the absence of C1 inhibitor. E. coli J-5 activated C1 about 10-fold more rapidly and bound approximately threefold more C1 than the other strains. E. coli J-5, but not the other strains, also bound C1s2, provided that the subcomponent was offered to the bacteria in the presence of C1q and calcium; such binding was thus independent of the presence or absence of C1r2. After C1 activation in the absence of C1 inhibitor, activated C1s spontaneously dissociated from E. coli 0111:B4, (P), and (A), but remained associated with E. coli J-5. The regulatory protein C1 inhibitor prevented C1 activation by the weaker activators, E. coli strains 0111:B4, (P), and (A), but had no effect on C1 activation by E. coli J-5. Although C1 inhibitor thus failed to modulate C1 activation by E. coli J-5, it did block the enzymatic activity of activated C1 bound to this strain. Analyses of the molecular processes involved revealed differences with other systems. In the presence of C1 inhibitor, the C1s subunit of C1 activated by E. coli J-5 underwent further cleavage with the release into the supernatant of C1s fragments and complexes of C1 inhibitor with light chain fragments. Such fragments were not disulfide-linked to the remainder of the C1s molecule. The bulk of the heavy chain remained adherent to the surface of E. coli J-5. This finding documents the presence of a binding site for activated C1s on the surface of E. coli J-5 and localizes this site to the heavy chain. These studies thus indicate that several E. coli strains are direct C1 activators. Furthermore, E. coli J-5 provides another example of a direct C1 activator having binding sites not only for C1q but also for dimeric C1s. The studies also show that there are multiple properties of particles which determine the ability to activate C1, the rate of activation, the possibility of regulation of the activation process by C1 inhibitor, and the fate of activated C1.