Sandbox Reserved 895: Difference between revisions

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==== '''[1.2.2.1] Overall Structural Analysis of RPE65''' ====

Using the crystal structure of bovine RPE65 (PDB: ''3FSN''), which is 99% similar to human RPE65, (although the crystal structure for human RPE65 is not currently available) as the basis of studying the RPE65 structure, RPE65 resembles a seven-bladed β-propeller with single-stranded extension on blades VI and VII and two-stranded extension on blade III shown in '''Figure 4'''. The top face of the β-propeller is defined by connecting the outer strand of the β-sheet with the inner strand of the next β-sheet. The iron cofactor is located near the top surface of the propeller which is coordinated by four His residues and three secondary Glu residues. Each blade of the propeller contributes one His residue to coordinate with the iron ion. A hydrophobic tunnel leads from the protein exterior to the active site which is defined by the iron ion to accommodate the passage of retinoids (which are conjugated to a fatty acid tail) from the membrane to the RPE65 catalytic site. The mouth of the tunnel is surrounded by three groups of nonpolar residues that contribute to the overall hydrophobicity of the tunnel and the integration with the lipid bilayer. There are also a few aromatic amino acid side chains that reside in this portion of the enzyme. This suggest that the depth of the RPE65 membrane is restricted to the proximal portions of the phospholipid acyl chains with respect to the polar head groups. Arg and Lys residues within this region also contribute to the association with the negatively charged phospholipid head groups. Using a helical wheel plot, Kiser and colleagues were able to determine that the tunnel region of RPE65 is amphipathic with positive charged residues separating the hydrophobic face from the hydrophilic face as shown in '''Figure 4C'''. <ref> DOI 19805034 </ref>

Using the crystal structure of bovine RPE65 (PDB: ''3FSN''), which is 99% similar to human RPE65, (although the crystal structure for human RPE65 is not currently available) as the basis of studying the RPE65 structure, RPE65 resembles a seven-bladed β-propeller with single-stranded extension on blades VI and VII and two-stranded extension on blade III shown in '''Figure 4'''. The top face of the β-propeller is defined by connecting the outer strand of the β-sheet with the inner strand of the next β-sheet. The iron cofactor is located near the top surface of the propeller which is coordinated by four His residues and three secondary Glu residues. Each blade of the propeller contributes one His residue to coordinate with the iron ion. A hydrophobic tunnel leads from the protein exterior to the active site which is defined by the iron ion to accommodate the passage of retinoids (which are conjugated to a fatty acid tail) from the membrane to the RPE65 catalytic site. The mouth of the tunnel is surrounded by three groups of nonpolar residues that contribute to the overall hydrophobicity of the tunnel and the integration with the lipid bilayer. There are also a few aromatic amino acid side chains that reside in this portion of the enzyme. This suggest that the depth of the RPE65 membrane is restricted to the proximal portions of the phospholipid acyl chains with respect to the polar head groups. Arg and Lys residues within this region also contribute to the association with the negatively charged phospholipid head groups. <ref> DOI 19805034 </ref>

[[Image:RPE65_Figure4_S7_blades.jpg|thumb|center|512 px|alt=Figure 4: RPE65 structure| '''Figure 4:''' Crystal structure of bovine RPE65 viewed from the bottom face of the seven-bladed β-propeller labeled in roman numerals from I to VII <ref> DOI 19805034 </ref>]]

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 ==== '''[1.2.2.1] Overall Structural Analysis of RPE65''' ====
-Using the crystal structure of bovine RPE65 (PDB: ''3FSN''), which is 99% similar to human RPE65, (although the crystal structure for human RPE65 is not currently available) as the basis of studying the RPE65 structure, RPE65 resembles a seven-bladed β-propeller with single-stranded extension on blades VI and VII and two-stranded extension on blade III shown in '''Figure 4'''. The top face of the β-propeller is defined by connecting the outer strand of the β-sheet with the inner strand of the next β-sheet. The iron cofactor is located near the top surface of the propeller which is coordinated by four His residues and three secondary Glu residues. Each blade of the propeller contributes one His residue to coordinate with the iron ion. A hydrophobic tunnel leads from the protein exterior to the active site which is defined by the iron ion to accommodate the passage of retinoids (which are conjugated to a fatty acid tail) from the membrane to the RPE65 catalytic site. The mouth of the tunnel is surrounded by three groups of nonpolar residues that contribute to the overall hydrophobicity of the tunnel and the integration with the lipid bilayer. There are also a few aromatic amino acid side chains that reside in this portion of the enzyme. This suggest that the depth of the RPE65 membrane is restricted to the proximal portions of the phospholipid acyl chains with respect to the polar head groups. Arg and Lys residues within this region also contribute to the association with the negatively charged phospholipid head groups. Using a helical wheel plot, Kiser and colleagues were able to determine that the tunnel region of RPE65 is amphipathic with positive charged residues separating the hydrophobic face from the hydrophilic face as shown in '''Figure 4C'''. <ref> DOI 19805034 </ref>
+Using the crystal structure of bovine RPE65 (PDB: ''3FSN''), which is 99% similar to human RPE65, (although the crystal structure for human RPE65 is not currently available) as the basis of studying the RPE65 structure, RPE65 resembles a seven-bladed β-propeller with single-stranded extension on blades VI and VII and two-stranded extension on blade III shown in '''Figure 4'''. The top face of the β-propeller is defined by connecting the outer strand of the β-sheet with the inner strand of the next β-sheet. The iron cofactor is located near the top surface of the propeller which is coordinated by four His residues and three secondary Glu residues. Each blade of the propeller contributes one His residue to coordinate with the iron ion. A hydrophobic tunnel leads from the protein exterior to the active site which is defined by the iron ion to accommodate the passage of retinoids (which are conjugated to a fatty acid tail) from the membrane to the RPE65 catalytic site. The mouth of the tunnel is surrounded by three groups of nonpolar residues that contribute to the overall hydrophobicity of the tunnel and the integration with the lipid bilayer. There are also a few aromatic amino acid side chains that reside in this portion of the enzyme. This suggest that the depth of the RPE65 membrane is restricted to the proximal portions of the phospholipid acyl chains with respect to the polar head groups. Arg and Lys residues within this region also contribute to the association with the negatively charged phospholipid head groups. <ref> DOI 19805034 </ref>
 [[Image:RPE65_Figure4_S7_blades.jpg|thumb|center|512 px|alt=Figure 4: RPE65 structure| '''Figure 4:''' Crystal structure of bovine RPE65 viewed from the bottom face of the seven-bladed β-propeller labeled in roman numerals from I to VII <ref> DOI 19805034 </ref>]]