Sandbox Reserved 768: Difference between revisions
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Although the full-length structure of mammalian PAH has not yet been clearly known, a large part of it has been identified. <ref name= "flydal"/>. It was solved by means of crystallizing the protein (at pH=7) to perform X-ray diffraction using molecular replacement. The search model used was based on the crystal structure of tyrosine hydroxylase because of the similarity between the two enzymes. <ref name="fusetti"> Fusetti, Fabrizia, Heidi Erlandsen, Torgeir Flatmark, and Raymond Stevens. "Structure of Tetrameric Human Phenylalanine Hydroxylase and Its Implications for Phenylketonuria." The Journal of Biological Chemistry 273.27 (1998): 16962-16967. Web. </ref>. The R-factor recorded was 0.251 and the mean B (or temperature) value was 33.0. <ref name= "pdb"/>. | Although the full-length structure of mammalian PAH has not yet been clearly known, a large part of it has been identified. <ref name= "flydal"/>. It was solved by means of crystallizing the protein (at pH=7) to perform X-ray diffraction using molecular replacement. The search model used was based on the crystal structure of tyrosine hydroxylase because of the similarity between the two enzymes. <ref name="fusetti"> Fusetti, Fabrizia, Heidi Erlandsen, Torgeir Flatmark, and Raymond Stevens. "Structure of Tetrameric Human Phenylalanine Hydroxylase and Its Implications for Phenylketonuria." The Journal of Biological Chemistry 273.27 (1998): 16962-16967. Web. </ref>. The R-factor recorded was 0.251 and the mean B (or temperature) value was 33.0. <ref name= "pdb"/>. | ||
'''Structure Revealed''' | '''Structure Revealed''' | ||
The monomeric unit of PAH is composed of three sites: an N-terminal regulatory | The monomeric unit of PAH is composed of three sites: an N-terminal regulatory | ||
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The catalytic domain contains the active site of the enzyme. It is composed of 13 α-helices and 8 β-strands <ref name= "fusetti"/> and houses the binding sites for the nonheme iron atom, the cofactor, and substrate. The iron binds to two histidines (His285 and His290 in hPAH) and a glutamate (Glu330) in the deep cleft in the core of each monomer. <ref name= "flydal"/>. | The catalytic domain contains the active site of the enzyme. It is composed of 13 α-helices and 8 β-strands <ref name= "fusetti"/> and houses the binding sites for the nonheme iron atom, the cofactor, and substrate. The iron binds to two histidines (His285 and His290 in hPAH) and a glutamate (Glu330) in the deep cleft in the core of each monomer. <ref name= "flydal"/>. | ||
The C-terminal oligomerization or tetramerization domain begins with an antiparallel-sheet | The C-terminal oligomerization or tetramerization domain begins with an antiparallel-sheet | ||
(residues 411–414, 421–424) | (residues 411–414, 421–424) | ||
and is formed by a C-terminal “arm” consisting of two β-strands, forming a β-ribbon, and a 40 Å long α-helix. This C-terminal arm extends over an adjacent monomer, thus bringing the four helices (one from each monomer) into a closely packed anti-parallel coiled-coil motif in the center of the structure (as can be seen in the tetramer structure above). <ref name= "fusetti"/>. The assembly of the enzyme occurs through a swapping mechanism in which the secondary structural elements mutually switch their position to promote oligomerization. <ref name= "fusetti"/>. | and is formed by a C-terminal “arm” consisting of two β-strands, forming a β-ribbon, and a 40 Å long α-helix. This C-terminal arm extends over an adjacent monomer, thus bringing the four helices (one from each monomer) into a closely packed anti-parallel coiled-coil motif in the center of the structure (as can be seen in the tetramer structure above). <ref name= "fusetti"/>. The assembly of the enzyme occurs through a swapping mechanism in which the secondary structural elements mutually switch their position to promote oligomerization. <ref name= "fusetti"/>. | ||
== PAH Mutations == | |||
Mutations and changes in the β-ribbon region have major detrimental effects on the enzyme stability. The mutations that lead | |||
to Phenylketonuria are most likely due to mutations in the PAH gene that cause changes at the junction between the catalytic and tetramerization domains of the enzyme. This leads to loss of enzyme stability and extensive misfolding which subsequently results in enzyme malfunction. The monomeric image below highlights the regions in the active site that, if mutated, would destroy enzyme activity and lead to phenylketonuria. | |||
[[Image:mutation sites.jpg]] | |||
The yellow atom is iron, the green structure is the BH4 cofactor, and the red sites are a few examples of mutation regions. | |||
