DHPR

Dihydropyridine receptor (DHPR) is a member of voltage-dqiendent Ca2+ channels (CaVi, L-type), which specifically binds to dihydropyridine derivatives, a group of the Ca2+ channel blockers. Cav 1.1 works as the voltage sensor for skeletal muscle contraction, and Cay 1.2, as Ca2+-influx channel for cardiac muscle contraction. 

In the skeletal muscle, RyRl can also mediate CICR. Therefore, it may reasonably be hypothesized that Ca2+ released from the RyR coupled with DHPR might... 

Figure 49-9. Possible chain of events leading to opening of the Ca release channel. As indicated in the text, the Ca voltage channel and the Ca release channel have been shown to interact with each other in vitro via specific regions in their polypeptide chains. (DHPR, dihydropyridine receptor RYR1, ryanodine receptor 1.)...

Figure 49-10. Simplified scheme of the causation of malignant hyperthermia (MIM 145600). At least 17 different point mutations have been detected in the RYRl gene, some of which are associated with central core disease (MIM 117000). It is estimated that at least 50% of families with members who have malignant hyperthermia are linked to the RYRl gene. Some individuals with mutations in the gene encoding DHPR have also been detected it is possible that mutations in other genes for proteins involved in certain aspects of muscle metabolism will also be found.

Hypokalemic periodic paralysis (MIM 114208) Slow Ca" voltage channel (DHPR) Skeletal muscle... 

Rarely, phenylketonuria results from a defect in the metabolism of biopterin, a cofactor for the phenylalanine hydroxylase pathway. The electron donor for phenylalanine hydroxylase is tetrahydrobiopterin (BH4), which transfers electrons to molecular oxygen to form tyrosine and dihydrobiopterin (QH2 Fig. 40-2 reaction 2). BH4 is regenerated from QH2 in an NADH-dependent reaction that is catalyzed by dihydropteridine reductase (DHPR), which is widely distributed. In the brain, this... 

Patients sustain convulsions and neurological deterioration. The urine contains low levels of the metabolites of serotonin, norepinephrine and dopamine. The reductase also plays a role in the maintenance of tetrahydrofolate levels in brain, and some patients have had low folate levels in the serum and CNS. Treatment has been attempted with tryptophan and carbidopa to improve serotonin homeostasis and with folinic acid to replete diminished stores of reduced folic acid. This therapy is sometimes effective. Diagnosis involves assay of DHPR in skin fibroblasts or amniotic cells. Phenylalanine hydroxylase activity is normal. 

DHPR dihydropteridine reductase GIRK G-protein-coupled inwardly rectifying K+ channel... 

All NMR experiments were performed on a Bruker DRX700 spectrometer operating at 700 MHz H frequency and equipped with a triple resonance probe and a triple axis gradient coil. DHPR concentration was 75 pM (300 mM monomer) in 25 pM d1rTris buffer, pH = 7.8 and 7=303 K. (Taken from Ref. [4]). 

The three-dimensional X-ray structure of the enzyme [19] reveals that several Thr residues occur in both the NADH cofactor and substrate binding sites (Fig. 21.5 A see p. 463). A Met residue (Metl7) is also present at the interface between the cofactor NADH and a substrate analog pyridine-2,6-dicarboxylate (PDC) (Fig. 21.5 A). Therefore, we prepared a sample of DHPR that was selectively labeled in these amino acid residues as follows 13C /1H Met, 13C /1H lie, 13C/1H Thr and uniformly 2H-labeled elsewhere ([MIT]-DHPR). This labeling can be achieved by supplementing the media with appropriate commercially available labeled amino acids, 12C/2H-labeled glucose and DzO [20] (see also the caption to Fig. 21.5 for details). 

Fig. 21.6 Establishing ligand binding with a cross-saturation experiment. A Chemical structure of NMNH. B Reference spectrum (blue) and binding site-saturated spectrum (red) of NMNH (500 pM) in the presence of [MITJ-DHPR at 2.5 pM...

Fig. 21.7 A Portion of a 2D [nH, H]-NOESY spectrum of NADH (500 pM) in complex with [MIT]-DHPR (75 pM) and PDC (500 pM). Protein-NADH NOEs are labeled with dashed circles. PDC-protein NOEs are labeled with solid circles. Inter-ligand PDC-NADH NOEs are labeled with dotted circles. Typical 2D [nH, HJ-NOESY [33] spectra were acquired with 256x2048 complex points and with mixing times between 50 ms and 500 ms. The measuring time for a 2D [nH, 1H]-...

BH4 deficiency can be caused by mutations in genes encoding the enzymes involved in its biosynthesis (GTPCH, PTPS, and SR) or regeneration (PCD/DCoH and DHPR). BH4 deficiency due to autosomal recessive mutations in BH4-metabo-lizing enzymes (except SR) has been described as a cause of hyperphenylalaninemia (HPA) [6]. Biochemical, clinical, and DNA data of patients with BH4 deficiencies are tabulated in the BIODEF and BIOMDB databases and are available on the Internet (www.bh4.org) [7]. Depending on the enzyme defect and the mode of inheritance,... 

Screening for a BH4 deficiency should be done in all newborns with plasma phenylalanine levels higher than 120 pmol/1, as well as in older children with neurologic signs and symptoms [10]. The following tests are recommended (1) analysis of pterins in urine, (2) measurement of DHPR activity in blood from a Guthrie card,... 

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... 

A Fig. 6.1.7a-k (continued) h control blood i blood PTPS deficiency j blood DHPR deficiency blood GTPCH deficiency. Neo Neopterin, Pri primapterin... 

Figure 6.1.9b,c shows chromatograms of reduced (electrochemical detection) and oxidized (fluorescence detection) pterins in the CSF of a 2-year-old control subject and a patient with DHPR deficiency. Elevated BH2 is characteristic for CSF from patients with DHPR and SR deficiency. In both groups BH4 concentrations are subnormal and oxidized biopterin is elevated. 

C DHPR-deficient CSF in a 6-month-old male. Note the elevated BH2. The shoulder on the peak is biopterin. Also note that BH4 concentration can be within the appropriate reference range in DHPR deficiency states [39]. The arrow denotes the injection point. Figure courtesy of Dr. Simon Heales, Fondon... 

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

Additional freeze-thawing of tissue or fibroblast lysates may result in reduced GTPCH activity, and it is thus recommended to assay activity always from freshly lysed material. Alternatively, lysates may be kept for 1-2 days at -80°C (this may not be the case for PTPS, SR, and DHPR assays, as extracts can be kept at -80°C for a longer period without loosing activity). 

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