Cyclohydrolase

In E. coli GTP cyclohydrolase catalyzes the conversion of GTP (33) into 7,8-dihydroneoptetin triphosphate (34) via a three-step sequence. Hydrolysis of the triphosphate group of (34) is achieved by a nonspecific pyrophosphatase to afford dihydroneopterin (35) (65). The free alcohol (36) is obtained by the removal of residual phosphate by an unknown phosphomonoesterase. The dihydroneoptetin undergoes a retro-aldol reaction with the elimination of a hydroxy acetaldehyde moiety. Addition of a pyrophosphate group affords hydroxymethyl-7,8-dihydroptetin pyrophosphate (37). Dihydropteroate synthase catalyzes the condensation of hydroxymethyl-7,8-dihydropteroate pyrophosphate with PABA to furnish 7,8-dihydropteroate (38). Finally, L-glutamic acid is condensed with 7,8-dihydropteroate in the presence of dihydrofolate synthetase. 

This interesting conversion of a five- into a six-membered heterocyclic ring was proven by the isolation of the enzyme GTP-cyclohydrolase from E. coli (71MI21600) and a similar one from Lactobacillus platarum (B-71MI21601) which catalyzes the reaction (300)(303). Dephosphorylation leads to 7,8-dihydro-D-neopterin (304), which is then cleaved in the side-chain to 6-hydroxymethyl-7,8-dihydropterin (305), the direct precursor of 7,8-dihy-dropteroic acid and 7,8-dihydrofolic acid (224). The alcohol (305) requires ATP and Mg " for the condensation with p-aminobenzoic and p-aminobenzoylglutamic acid, indicating pyrophosphate formation to (306) prior to the substitution step. 

Determination of GTP cyclohydrolase and D-erythro-7,8-dihydroneopterin triphosphate synthetase... 

FIGURE 40-2 The phenylalanine hydroxylase (PAH) pathway. Phenylketonuria usually is caused by a congenital deficiency of PAH (reaction 1), but it also can result from defects in the metabolism of biopterin, which is a cofactor for the hydroxylase. Enzymes (1) Phenylalanine hydroxylase (2) Dihydropteridine reductase (3) GTP cyclohydrolase (4) 6-pyruvoyltetrahydrobiopterin synthase. BH4, tetrahydrobiopterin DEDT, o-erythro-dihydroneopterin triphosphate QH2, dihydrobiopterin. 

Other causes of PKU secondary to defective tetrahydrobiopterin synthesis include GTP cyclohydrolase deficiency and 6-pyravoyltetrahydrobiopterin synthase deficiency. Patients with either defect have psychomotor retardation, truncal hypotonia with limb hypertonia, seizures and a tendency to hyperthermia. The intravenous administration of BH4 may lower blood phenylalanine levels but this cofactor may not readily cross the blood-brain barrier. Treatment with synthetic pterin analogs or supplementation with tryptophan and carbidopa may prove more efficacious, particularly if treatment is started early in life. 

These patients suffer from a genetic defect of dopamine synthesis, caused by reduced GTP cyclohydrolase activity. This enzyme is rate-limiting in the biosynthesis of tetra-hydrobiopterin, a cofactor of the dopamine-synthesizing enzyme tyrosine hydroxylase (see Fig. 40-2). 

Grabarse W, Vaupel M, Vorholt JA, et al. 1999. The crystal structure of methenyl-tetrahydromethanopterin cyclohydrolase from the hyperthermophilic archaeon Methanopyrus kandleri. Structure Fold Des 7 1257-68. 

Vorholt JA, Chistoserdova L, Stolyar SM, et al. 1999. Distribution of tetrahy-dromethanopterin-dependent enzymes in methylotrophic bacteria and phylogeny of methenyl tetrahydromethanopterin cyclohydrolases. J Bacteriol 181 5750-7. 

Pomper BK, Vorholt JA, Chistoserdova L, et al. 1999. A methenyl tetrahydromethanopterin cyclohydrolase and a methenyl tetrahydrofolate cyclohydrolase in Methylobacterium extorquens AMI. Eur J Biochem 261 475-80. 

This group includes the coenzyme forms of water-soluble vitamin B2 or riboflavin. Synthesis occurs by initial cyclohydrolase action on the guanine ring of GTP and subsequent steps lead to the synthesis of the isoalloxazine ring structure (see structures below). 

This enzyme [EC 1.1.1.23] catalyzes the reaction of l-histidinol with two NAD+ to produce L-histidine and two NADH. L-Histidinal will also serve as a substrate for this protein. The Neurospora crassa enzyme will also catalyze the reactions of phosphoribosyl-AMP cyclohydrolase [EC 3.5.4.19] and phosphoribosyl-ATP pyrophosphatase [EC 3.6.1.31]. 

This enzyme [EC 3.6.1.31] catalyzes the hydrolysis of 5-phosphoribosyl-ATP to produce 5-phosphoribosyl-AMP and pyrophosphate (or, diphosphate). The Neurospora crassa enzyme also catalyzes the reactions of histidinol dehydrogenase and phosphoribosyl-AMP cyclohydrolase. 

GTP CYCLOHYDROLASES GUANYLATE CYCASE HEXOSE-1-PHOSPHATE GUANYLYL-TRANSFERASE... 

CMP SYNTHETASE GTP CYCLOHYDROLASE II GUANYLATE CYCLASE HEXOSE-1-PHOSPHATE CUANYLYL-TRANSEERASE... 

The enzymes in the zebra fish pathway are presumably very similar to those of other vertebrates. However a completely different type of GTP cyclohydrolase has been identified in the hyperthermophilic euryarchaeon, Methanococcus jannashii <2002B15074>. This enzyme, in purified recombinant form, produced as a stable end product 2-amino-5-formylamino-6-ribofuranosylamino-4(3//)-pyrimidinone monophosphate, a compound that is an intermediate in the action of normal GTP cyclohydrolases. The biosynthesis of the incorporation of the pterin into methanopterin in Methanobacterium thermoautotrophicum has been proposed to occur via substitution of 7,8-dihydro-6-hydroxymethylpterin diphosphate 227 (Scheme 44) <1998BBA257>. 

An important discovery is that procaryotes contain a different GTP cyclohydrolase 1 family distinct from the well studied canonical eukaryotic enzyme. Potentially, this enzyme is a target for new antibacterial drugs <2006JBC37586>. 

A Fig. 6.1.7a- HPLC of pterins using a column-switching system a standard mixture b control urine c urine guanosine triphosphate cyclohydrolase I (GTPCH) deficiency d urine 6-pyru-voyl-tetrahydropterin synthase (PTPS) deficiency e urine pterin-4a-carbinolamine dehydratase (PCD) deficiency f urine dihydropteridine reductase (DHPR) deficiency g urine phenylketonuria 4-8 h after tetrahydrobiopterin (BH4) administration h-k see next page... 

DHPR Dihydropteridine reductase, DRD dopa-responsive dystonia, GTPCH GTP cyclohydrolase I, n normal, PCD pterin-4a-carbinolamine dehydratase, PTPS 6-pyruvoyltetrahydrobiopterin synthase, SR sepiapterin... 

To prepare lysates from (nonstimulated) fibroblasts, cells from one confluent 78-cm2 plate are suspended in 0.15 ml lysis buffer (see below) and lysed by freezing and thawing six times and subsequent centrifugation at 13,000 x for 5 min. An aliquot of 0.05 ml of the supernatant is directly used for the enzyme assay. The preparation of tissue homogenate is described in section 6.1.4.1. GTP cyclohydrolase I, subheading Specimen . 

Ichinose H, Ohye T, Takahashi E, Seki N, Hori T, Segawa M, Nomura Y, Endo K, Tanaka H, Tsuji S, Fujita K, Nagatsu T (1994) Hereditary progressive dystonia with marked diurnal fluctuation caused by mutation in the GTP cyclohydrolase I gene. Nature Genet 8 236-241... 

Viveros OH, Lee CL, Abou-Donia MM, Nixon JC, Nichol CA (1981) Biopterin cofactor biosynthesis independent regulation of GTP cyclohydrolase in adrenal medulla and cortex. Science 213 349-350... 

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