Preparation of 5‐amino‐4‐imidazole‐N‐succinocarboxamide ribotide, 5‐amino‐4‐imidazole‐N‐succinocarboxamide riboside and succinyladenosine, compounds usable in diagnosis and research of adenylosuccinate lyase deficiency

Summary The enzyme adenylosuccinate lyase (ADSL) intervenes twice in the biosynthesis of adenine nucleotides. ADSL deficiency is an inherited metabolic disease characterized by various degrees of psychomotor retardation and accumulation of dephosphorylated enzyme substrates 5‐amino‐4‐imidazole‐ N ‐succinocarboxamide riboside (SAICAr) and succinyladenosine (SAdo) in body fluids. Severity of symptoms seems to correlate with residual activity of mutant enzyme and with SAdo/SAICAr concentration ratio in cerebrospinal fluid. To better understand the pathogenetic mechanisms of the disease symptoms, studies of catalytic properties of mutant enzymes together with in vitro and in vivo experiments utilizing SAICAr and SAdo must be performed. Such studies require availability of both ADSL substrates, 5‐amino‐4‐imidazole‐ N ‐succinocarboxamide ribotide (SAICAR) and succinyladenosine 5′‐monophosphate (SAMP) and their dephosphorylated products in sufficient amounts and purity. Except for SAMP, none of these compounds is commercially available and they must therefore be synthesized. SAICAR was prepared by recombinant human ADSL‐catalysed reaction of AICAR (5‐aminoimidazole‐4‐carboxamide) with fumarate and isolated by thin‐layer chromatography. SAICAr and SAdo were prepared by calf intestine alkaline phosphatase‐catalysed dephosphorylation of SAICAR and SAMP and isolated on cation‐ and anion‐exchange resin columns. The procedures described are easily scalable and provide high yields of sufficiently pure products for use in experiments related to studies of pathogenetic mechanisms in ADSL deficiency.