RESUMEN
Flow dialysis can be used to measure (i) ligand binding to macromolecules and (ii) the size of transmembrane ion gradients. Generally an approximate method is used to calculate the binding or gradient parameters from the raw data. Here we present a simple but exact method and evaluate the errors that may arise when the approximate method is used to calculate the magnitude of ion gradients. In addition, equations are presented that allow for a correction for sampling from or additions to the upper compartment of a flow-dialysis vessel during the measurements. Setty and Hendler [(1982) J. Biochem. Biophys. Methods 7, 35-46] have reported artifacts in the measurement of ion-gradients caused by the addition of electron donors to the upper compartment of a flow-dialysis cell. Here we extend their observations and suggest additional methods to prevent such artifacts.
Asunto(s)
Biopolímeros , Diálisis/métodos , Sustancias Macromoleculares , Ligandos , Modelos TeóricosRESUMEN
Oxygen consumption following isometric tetanic contractions of single fibres and multifibre preparations of the tibialis anterior muscle of Rana temporaria was determined by continuous polarographic measurement of the PO2 in a 280 microliter glass chamber. Mixing of the fluid surrounding the muscle was achieved by an Archimedian screw. Force was measured via a stainless-steel wire leaving the chamber via a glass capillary. The characteristics of the oxygen-measuring system were assessed by injection of 1.6 microliter dye into the chamber and filming its subsequent distribution, and by injection of 1.6 microliter Ringer solution with a high (or low) oxygen content into the chamber and measuring the subsequent change of oxygen. It was found that a change in oxygen was measured after a true delay of 3 s and with an over-all time constant of 3.25 s following that delay. For seven single fibres the oxygen consumption following a 3 s tetanus was on average 2.46 mumol g-1; the average integrated value of the developed stress was 0.98 N mm-2 s. These two values were on average about 45% lower for the same tetani of multifibre preparations, but the average ratio of oxygen consumption to integrated stress was the same. Oxygen consumption was varied by changing tetanus duration. When the amount of oxygen consumed was plotted against stress integral a non-linear relationship was found because oxygen consumption increased less than the integrated stress value with longer tetani. Oxygen consumption did not start at the onset of contraction but about 10 s later. It then followed an exponential time course with an average time constant of 120 s. Delay and time constant were independent of the amount of oxygen consumed. The finding that oxygen consumption follows contraction after a delay of a few seconds confirms a similar conclusion drawn indirectly from studies on recovery heat by other investigators. A dependency of the time course of oxygen consumption on tetanus duration, as reported in the literature for frog muscle at 0 degree C, was not found.
Asunto(s)
Contracción Muscular , Músculos/metabolismo , Consumo de Oxígeno , Animales , Técnicas In Vitro , Rana temporaria , Estrés Mecánico , Temperatura , Factores de TiempoAsunto(s)
Miocardio/metabolismo , Consumo de Oxígeno , Animales , Perros , Matemática , Modelos BiológicosAsunto(s)
Metabolismo Energético , Mitocondrias Hepáticas/metabolismo , Adenosina Difosfato/metabolismo , Adenosina Trifosfato/metabolismo , Animales , Transporte de Electrón , Glutamatos/metabolismo , Concentración de Iones de Hidrógeno , Hipertiroidismo/metabolismo , Membranas/metabolismo , Fosfatos/metabolismo , Ratas , Succinatos/metabolismo , TermodinámicaAsunto(s)
Membranas Artificiales , Mitocondrias Hepáticas/metabolismo , Desacopladores/farmacología , Animales , Colesterol , Cromatografía en Capa Delgada , Yema de Huevo , Femenino , Ferricianuros/metabolismo , Liposomas , Hígado/citología , Fosforilación Oxidativa , Consumo de Oxígeno , Fosfatidilcolinas , Ratas , EspectrofotometríaRESUMEN
Evidence is presented that progametangia in both the plus and the minus mating types of Mucor mucedo can be induced by one substance, namely (-)-trisporic acid B. A method is described for the determination of the concentration of the sex factors (trisporone, trisporic acid B, trisporic acid C) in mated cultures of Mucorales by polarography. It can be demonstrated that the amount of plus mycelium is limiting for the production of the sex factors in Blakeslea trispora. It is shown that the minus type of this organism is able to synthesize the sex factors when incubated in the filtered medium of a mated culture. Cycloheximide and 5-fluorouracil inhibit strongly the sex factor production in a mated culture of B. trispora at any time. This result suggests that sexual activity comprises the synthesis of proteins which are involved in the production of the sex factors.