Stimulation of rat liver mitochondrial adenosine triphosphatase by anions.

The hydrolysis of MgATP by isolated rat liver mitochondrial ATPase (EC 3.6.1.3) at pH 8.0 was stimulated by various anions. The rate of hydrolysis was increased from 18 to 170 mumol per min per mg, a 9.4-fold stimulation, by HSeO3 at 1 mM MgATP. In the absence of a stimulatory anion, reciprocal plots of initial velocity studies with MgATP as the variable substrate were curved (Hill coefficient approximately 0.5). With the addition of anion, the reciprocal plots became linear. When the substrate was MgITP or MgGTP with the isolated enzyme or MgATP with submitochondrial particles, no curvature of the reciprocal plots was observed. With purified ATPase, anions stimulated the hydrolysis of MgITP, MgGTP, MgUTP or MgCTP only slightly. With submitochondrial particles the stimulation by anions of MgATP hydrolysis was limited to approximately 2-fold. These data are interpreted to indicate the existence of two substrate sites for MgATP and an anion-binding site on the isolated enzyme.     

The influence of ammonium and calcium lons on gluconeogenesis in isolated rat hepatocytes and their response to glucagon and epinephrine

The use of n-butylmalonate as an inhibitor of malate transport from mitochondria and of aminooxyacetate as an inhibitor of glutamate-aspartate transaminase indicated that rat liver hepatocytes employ the aspartate shuttle for gluconeogenesis from lactate which supplies reducing equivalents to the cytosolic NAD system. In contrast, malate is transported from mitochondria to cytosol for gluconeogenesis from pyruvate. This conclusion is corroborated by the finding that the addition of ammonium ions enhances gluconeogenesis from lactate but inhibits glucose formation from pyruvate. In hepatocytes, glucagon and epinephrine have relatively little effect on glucose synthesis from lactate. Ammonium ions permit both of these hormones to exert their usual stimulation of gluconeogenesis from lactate.Calcium ions (1.3 mm) enhance gluconeogenesis from lactate and from lactatepyruvate mixtures (10:1). The stimulatory effects of Ca²⁺ and NH₄⁺ are additive and, when lactate is the substrate, the rates of gluconeogenesis achieved are so high as to preclude further stimulation by glucagon.     

Pyrrolo[1,2-b]pyridazin-2-ones as potent inhibitors of HCV NS5B polymerase

Pyrrolo[1,2-b]pyridazin-2-one analogs were discovered as a novel class of inhibitors of genotype 1 HCV NS5B polymerase. Structure-based design led to the discovery of compound 3 k, which displayed potent inhibitory activities in biochemical and replicon assays (IC(50) (1b)<10nM; EC(50) (1b)=12 nM) as well as good stability towards human liver microsomes (HLM t(1/2)>60 min).