Evidence for the hypothesis that 10-formyldihydrofolate is the in vivo substrate for aminoimidazolecarboxamide ribotide transformylase

Abstract

We postulate that 10-formyl-7,8-dihydrofolate (10-HCO-H(2)folate), not 10-formyl-5,6,7,8-tetrahydrofolate (10-HCO-H(4)folate), is the predominant in vivo substrate for mammalian aminoimidazolecarboxamide ribotide (AICAR) transformylase, an enzyme in purine nucleotide biosynthesis de novo, which introduces carbon 2 (C(2)) into the purine ring. 10-HCO-H(2)folate exists in vivo as labeled 10-formyl-folic acid (10-HCO-folic acid: an oxidation product of 10-HCO-H(4)folate and 10-HCO-H(2)folate) and is found after doses of labeled folic acid in humans or laboratory animals. The bioactivity of the unnatural isomer, [6R]-5-formyltetrahydrofolate, in humans is explained by its in vivo conversion to 10-HCO-H(2)folate. The structure and active site of AICAR transformylase are not consistent with other enzymes that utilize 10-HCO-H(4)folate. Because 10-HCO-H(4)folate is rapidly oxidized in vitro to 10-HCO-H(2)folate by cytochrome C alone and in mitochondria, it is hypothesized that this process takes place in vivo. In vitro data indicate that 10-HCO-H(2)folate is kinetically preferred over 10-HCO-H(4)folate by AICAR transformylase and that this enzyme may not have access to sufficient supplies of 10-HCO-H(4)folate. Methotrexate blockage of the AICAR transformylase process in patients with rheumatoid arthritis suggests that dihydrofolate (H(2)folate) reductase is involved and is consistent with H(2)folate and 10-HCO-H(2)folate being the product and substrate for AICAR transformylase. The labeling of purine C(2) by an oral dose of [6RS]-5-H[(13)C]O-H(4)folate in a human subject is consistent with 10-H[(13)C]O-H(2)folate formation from unnatural isomer, [6R]-5-H[(13)C]O-H(4)folate, and it being a substrate for AICAR transformylase. In vitro exchange reactions of purine C(2) using H(4)folate coenzymes are not duplicated in vivo and is consistent with H(2)folate coenzymes being used in vivo by AICAR transformylase.

A marriage of two “Methusalem” drugs for the treatment of psoriasis?

“…The importance of adenosine as an anti-inflammatory mediator was questioned 53 , 54 and could not been convincingly demonstrated in later experiments 55 . It is, however, generally accepted that AICAR (ZMP) increases after methotrexate 56 —either by direct inhibition of AICART or alternatively, favoring the reverse reaction by increasing dihydrofolate via blockade of dihydrofolate reductase 52 . (For more details, see the legend of Fig.…”

“…These are based on the reversibility of the reaction (indicated by the double-headed arrow) and that AICART has rather low affinity for the polyglutamates. As an exception to all other reactions in mammalian one-carbon metabolism 51 10-formyl-7, 8-dihydrofolate and dihydrofolate can be substrate and product, respectively 52 . Increase of the product by blockade of the dihydrofolate reductase will slow the reaction and lead to increase in AICAR.…”

Physiological levels of folic acid reveal purine alterations in Lesch-Nyhan disease

“…The authors hypothesized that it could be due to the differential binding mechanisms of the substrates to the enzymes: GART can bind its two substrates in a random manner (27), whereas ATIC obeys an ordered sequential binding mechanism, with 10-formyltetrahydrofolate binding first before ZMP can bind (28). Alternatively, it has been reported that human ATIC and GART do not utilize the same 10-formyltetrahydrofolate pool (29,30). Thus, ATIC can use 10-formyltetrahydrofolate formed from tetrahydrofolate and histidine or formate, whereas GAR transformylase, which is in a complex with the trifunctional folate-metabolizing enzyme, can use 10-formyltetrahydrofolate formed from tetrahydrofolate and glycine or serine.…”

Folate-Dependent Purine Nucleotide Biosynthesis in Humans

“…as a substrate in vitro (8,9); 3) the unnatural isomer, [6R]-5-formyltetrahydrofolate (5-formyl-H 4 folate), is inactive as a substrate for folate-metabolizing enzymes in vitro but bioactive in vivo in humans, suggesting its conversion to 10-formyl-H 2 folate in vivo (10); and 4) the kinetics of C 2 and C 8 enrichment of uric acid in humans by [6RS]5-13 C-formyl-H 4 folate are different, suggesting that GAR and AICAR transformylases do not utilize the same 10-formyl-H 4 folate pool (9). In this review, we discuss the distribution of onecarbon from ring-2-C of histidine, 2-C of glycine, 3-C of serine, and formate to C 2 and/or C 8 of uric acid as well as other purines.…”