Folylpoly-gamma-glutamates as cosubstrates of 10-formyltetrahydrofolate:5'-phosphoribosyl-5-amino-4-imidazolecarboxamide formyltransferase.

N10-Formyltetrahydropteroylpoly-gamma-glutamates (N10-formyl-H4PteGlun) having n = 1, 3, 4, 5, 6, and 7 glutamyl residues have been tested as cosubstrates of the purine biosynthesis enzyme 10-formyltetrahydrofolate:5'-phosphoribosyl-5-amino-4-imidazolecarboxamide formyltransferase (AICAR transformylase) of chicken liver. The cosubstrates were synthesized by solid-phase synthesis, reduced catalytically, and formylated; a purified enzyme preparation was used and assayed spectrophotometrically following the deltaOD at 298 nm resulting from conversion of the formylated folate to the free tetrahydro form. Km values and Vm values determined at saturating concentrations of AICAR and at 25 and 150 mM KC1 were used to calculate the relative specificity constants Vm/Km for the N10-formyl-H4PteGlun. At physiologic [K+] (150 mM) they were 1.0, 52, 250, 93, 120, and 59 and at the lower (25 mM) [K+] the relative specificity constants were 1.0, 64, 78,34, 48, and 37 for n = 1, 3, 4, 5, 6, and 7, respectively. The poly-gamma-glutamates are clearly the preferred cosubstrates, particularly when tested at physiologic [K+]. The maximal relative specificity constant observed with N10-formyl-H4PteGlu4 supports the hypothesis that regulation of certain pathways of one-carbon metabolism may operate via alterations of the poly-gamma-glutamyl chain length. No inhibition by the unnatural (d) isomers of the N10-formyl-H4PteGlun was observed.     

Protein kinase inhibitors block the stimulation of the AMP-activated protein kinase by 5-amino-4-imidazolecarboxamide riboside

The AMP-activated protein kinase (AMPK) is the central component of a protein kinase cascade that plays a major role in energy sensing. AMPK is activated pharmacologically by 5-amino-4-imidazolecarboxamide (AICA) riboside monophosphate (ZMP), which mimics the effects of AMP on the AMPK cascade. Here we show that uptake of AICA riboside into cells, mediated by the adenosine transport system, is blocked by a number of protein kinase inhibitors. Under these conditions, ZMP does not accumulate to sufficient levels to stimulate AMPK. Our results demonstrate that careful interpretation is required when using AICA riboside in conjunction with protein kinase inhibitors to investigate the physiological role of AMPK.     

The Accumulation of 5′-Phosphoribosyl-5-amino-4-imidazolecarboxamide in Folate-deficient Pea Seedlings and the Enzymatic Reaction in which the Compound is Involved

The functions of isoflavonoids and related compounds as well as of their degradation products in the allelopathy of red clover were investigated from chemical standpoint. Susceptibilities of red clover to isoflavonoids, related compounds and their degradation products were higher than those of various plants including white and alsike clovers, and contents of the isoflavonoids in red clover were extremely higher as compared with those in white clover and orchard grass. Inhibitory substances were isolated from the soil capable to cause “clover sickness,” but they were not isoflavonoids but phenolic acids considered to be originated probably in the formers. These acids were also obtained together with some kinds of isoflavonoids from the waste culture solution used for cultivation of red clover seedlings. Finally degradation process of isoflavonoids was followed up in neutral, acidic and alkaline solutions.