Sepiapterin Reductase Placemat Neurotransmitters Module: The Beery Twins’ Story© A Project-Based Learning Activity ...where molecules become real TM

I. The Beery Family History IV. Validation and Segregation of Two SPR Alleles VI. The Biosynthesis of Two Neurotramsmitters Guanosine Triphosphate (GTP) Figure 3. GCH1 Metabolic Soon after Alexis and Noah Beery were born it was obvious to The Sanger sequencing traces (below) show the SPR genotype for Pathways of Sepiapterin reductase is the final enzyme in the biosynthetic Dihydroneopterin Triphosphate their parents, Joe and Retta, that the twins were different from each member of the Beery family. Neurotransmitter pathway for – a cofactor used by other enzymes their older brother, Zach. They demonstrated poor muscle tetrahydrobiopterin PTPS Production1 in the synthesis of the neurotransmitters dopamine and serotonin. tone, cried nonstop, vomited frequently and missed The Arg150Gly mutation is an A G mutation on chromosome 2 at 6-Pyruvoyl-tetrahydropterin developmental milestones. Physicians diagnosed the twins nucleotide 72,969,094 leading to the replacement of Arginine with with cerebral palsy. By age 5, Alexis was having difficulty Glycine. The unaffected father is heterozygous (A/G) for the In the case of dopamine biosynthesis, the enzyme tyrosine SPR swallowing and was wasting away, symptoms not consistent pathogenic Arg150Gly allele at the first locus and homozygous (A/A) hydroxylase uses tetrahydrobiopterin as a cofactor to convert tyrosine Tetrahydrobiopterin (BH4) with cerebral palsy. Retta came across an article about a rare for the wild-type allele at the second locus. to L-DOPA. In the case of serotonin, the enzyme tryptophan disorder, dopa-responsive dystonia (DRD), which is caused by hydroxylase uses tetrahydrobiopterin to convert tryptophan to TH TPH a deficiency of the brain The Lys251X mutation is an A T mutation on chromosome 2 at 5-hydroxy tryptophan (5-HTP). In a second reaction in each pathway, Tyrosine Tryptophan PAH Phenylalanine neurotransmitter dopamine. nucleotide 72,972,139 aromatic L-amino acid decarboxylase (AAAD) converts both L-DOPA The symptoms matched those resulting in the to dopamine and 5-HTP to serotonin, the active neurotransmitters. L-DOPA 5-Hydroxtryptophan (5-HTP) Tyrosine of her daughter. Shortly after, conversion of a Lysine AAAD AAAD both twins were treated with codon (AAG) to a STOP Retta What can go wrong? L-DOPA (dopamine precursor), codon (UAG). The Dopamine Serotonin Mutations in the gene for sepiapterin reductase results in a and their condition improved unaffected mother is Joe dramatically. However, as the homozygous (A/A) for deficiency in the cofactor tetrahydrobiopterin – which in turn results in deficiencies in dopamine and serotonin. Enzymes Neurotransmitters Cofactors twins grew older, it became the wild-type allele at the Zach apparent that even this first locus but diagnosis was incorrect as their heterozygous (A/T) for How can we fix it? But Life is Complicated health began to further the pathogenic Lys251X Alexis Neurotransmitter levels can be boosted Aromatic L-amino acid decarboxylase is present in the bloodstream where it deteriorate. allele at the second Noah locus. in individuals with a defective sepiapterin rapidly converts the daily dose of L-DOPA to dopamine. While L-DOPA can reductase enzyme with daily doses of easily cross the blood/brain barrier, dopamine cannot. Therefore, to optimize the Each affected twin is a both L-DOPA and 5-HTP. Both of these delivery of L-DOPA to the brain where it is needed, the twins’ medication is Figure 2. Pedigree and validation compound heterozygote - precursors are downstream of the supplemented with carbidopa – an inhibitor of aromatic L-amino acid of two deleterious SPR Alleles1 II. The Beery Family Pedigree (A/G and A/T) with a biosynthetic step that is catalyzed by an decarboxylase. In the presence of carbidopa, the half-life of the L-DOPA in the enzyme that requires bloodstream is increased so that sufficient quantities cross the blood/brain Further examination of the Beery Family pedigree shows a different pathogenic tetrahydrobiopterin. barrier, where it is then decarboxylated to form the active neurotransmitter. family history of depression on the paternal side and a history mutation at each allele. of fibromyalgia and undisclosed neurological disorder on the maternal side, in addition to DRD in the twins.

Unaffected Based on 1sep.pdb D DRD Dystonia VII. The Beery Family Today! Other Neurological Disorder F D F Fibromyalgia Noah and Alexis have a passion D Depression to share their story. According to Deceased D Joe F Retta Miscarriage a recent blog post, ”It's our hope and prayer that people will use Male Female their voices in their medical care or any other area of their lives, Zach Alexis Noah and never, ever give up hope - no matter how dire the situation may Figure 1. Pedigree of a Family1 seem. Because hope is unlimited and has no boundaries!”

III. Whole Genome Sequencing References In the case of the Beery twins, whole genome sequencing led to a more complete understanding of the molecular basis 1. Gibbs et al. (2011). Whole-Genome Sequencing for of their disease and informed a change in their medical Optimized Patient Management. Science Translational treatment. Joe Beery was hired by Life Technologies (a Medicine. 3(87):1-6 biotech company involved in NextGen DNA sequencing) V. Mapping the Mutations to a Physical Model of Sepiapterin Reductase where he arranged to have the twins’ genomes sequenced 2. Huber et al. (1997). The 1.25 A crystal structure of by the Baylor College of Medicine.1 Various bioinformatic - Joe Beery’s Arg150Gly missense mutation (left image) results in a change from a positively charged basic arginine amino acid at position 150 sepiapterin reductase reveals it binding mode to pterins and filters were applied to eliminate variants that were not to an uncharged glycine. This would disrupt a salt bridge interaction with the negatively charged aspartic acid at 144 in the functional enzyme. brain neurotransmitters. The EMBO Journal. 16(24): thought to be responsible for their condition. Only three 7219-7230. variants remained as candidate genes. One of these was - Retta Beery’s Lys251X nonsense mutation (right image) results in a change from lysine at position 251 to a premature STOP codon, causing sepiapterin reductase (SPR). truncation of the enzyme. 3. http://dystonia.thebeerys.com Version 1.2 - 9/2015