The observed equilibrium constants (Kobs) of the creatine kinase (EC 2.7.3.2), myokinase (EC 2.7.4.3), glucose-6-phosphatase (EC 3.1.3.9), and fructose-1,6-diphosphatase (EC 3.1.3.11) reactions have been determined at 38 degrees C, pH 7.0, ionic strength 0.25, and varying free magnesium concentrations. The equilibrium constant (KCK) for the creatine kinase reaction defined as: KCK = [sigma ATP] [sigma creatine] divided by ([sigma ADP] [sigma creatine-P] [H+]) was measured at 0.25 ionic strength and 38 degrees C and was shown to vary with free [Mg2+]. The value was found to be 3.78 x 10(8) M-1 at free [Mg2+] = 0 and 1.66 x 10(9) M-1 at free [Mg2+] = 10(-3) M. Therefore, at pH 7.0, the value of Kobs, defined as Kobs = KCK[H+] = [sigma ATP] [sigma creatine] divided by ([sigma ADP] [sigma creatine-P] was 37.8 at free [Mg2+] = 0 and 166 at free [Mg2+] = 10(-3) M. The Kobs value for the myokinase reaction, 2 sigma ADP equilibrium sigma AMP + sigma ATP, was found to vary with free [Mg2+], being 0.391 at free [Mg2+] = 0 and 1.05 at free [Mg2+] = 10(-3) M. Taking the standard state of water to have activity equal to 1, the Kobs of glucose-6-P hydrolysis, sigma glucose-6-P + H2O equilibrium sigma glucose + sigma Pi, was found not to vary with free [Mg2+], being 110 M at both free [Mg2+] = 0 and free [Mg2+] = 10(-3) M. The Kobs of fructose-1,6-P2 hydrolysis, sigma fructose-1,6-P2 equilibrium sigma fructose-6-P + sigma Pi, was found to vary with free [Mg2+], being 272 M at free [Mg2+] = 0 and 174 M at free [Mg2+] = 0.89 x 10(-3) M.
It has been shown that endocytic vesicles in BALB/c 3T3 cells have a pH of 5.0 (Tycko and Maxfield, Cell, 28:643-651). In this paper, a method for measuring the effect of various agents, including weak bases and ionophores, on the pH of endocytic vesicles is presented. The method is based on the increase in fluorescein fluorescence with 490-nm excitation as the pH is raised above 5.0. Intensities of cells were measured using a microscope spectrofluorometer after internalization of fluorescein-labeled alpha 2-macroglobulin by receptor-mediated endocytosis. The increase in endocytic vesicle pH was determined from the increase in fluorescence after addition of various concentrations of the test agents. The following agents increased endocytic vesicle pH above 6.0 at the indicated concentrations: monensin (6 microM), FCCP (10 microM), chloroquine (140 microM), ammonia (5 mM), methylamine (10 mM). The ability of many of these agents to raise endocytic vesicle pH may account for many of their effects on receptor-mediated endocytosis. Dansylcadaverine caused no effect on vesicle pH at 1 mM. The observed increases in vesicle pH were rapid (1-2 min) and could be reversed by removal of the perturbant. This reversibility indicates that the vesicles themselves contain a mechanism for acidification. The increase in vesicle pH due to these treatments can be observed visually using an SIT video camera. Using this method, it is shown that endocytic vesicles become acidic at very early times (i.e., within 5-7 min of continuous uptake at 37 degrees C).
Abstract The paper comprises an (s,S) production inventory facility in which multiple vacations and varying service rates are considered for the multiple servers. The arrival of customers constitutes a Markovian Arrival Process (MAP) and service completion times follow the phase type distributions with representations ( α, S ) and ( β, T ) respectively. Manufacturing needs to begin at the moment when the stock position falls to s. Service is offered at a lower rate if the stock level ranges from 0 to s and the service time distribution has the representations (α, η 1 S) and (β, η 2 T ), 0 < η 1, η 2 < 1 respectively. If the stock level reaches the maximum S, production is ceased to operate. 1-limited service policy, Bernoulli service schedule, and Exhaustive service disciplines are considered for the servers. A suitable cost function is outlined as per performance assessment. The impact of negative correlated inter arrival times on cost function is illustrated. Also, a relative study of expected cost function on different service modes is presented.