Sandbox Reserved 822: Difference between revisions
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The standard PH domain fold consists of mainly three different sections: | The standard PH domain fold consists of mainly three different sections: | ||
*One section is a <scene name='56/568020/Barrel-like/3'>barrel-like structure</scene> formed by residues 456-530 (see Fig.1, green section). This structure is formed by two, almost orthogonal, β sheets, one consisting of four (β1 - β4) and one of three (β5 - β7) stand. | *One section is a <scene name='56/568020/Barrel-like/3'>barrel-like structure</scene> formed by residues 456-530 (see Fig.1, green coloured section). This structure is formed by two, almost orthogonal, β sheets, one consisting of four (β1 - β4) and one of three (β5 - β7) stand. | ||
*One side of the barrel is blocked by a <scene name='56/568020/Alpha1/1'>C-terminal alpha helix</scene> (see Fig.1, red section). | *One side of the barrel is blocked by a <scene name='56/568020/Alpha1/1'>C-terminal alpha helix</scene> (see Fig.1, red coloured section). | ||
*On the other side of the barrel three variable loops (VL 1-3) form a bowl-like structure which is lined by positively charged residues, consituting the phosphoinositide-binding site. | *On the other side of the barrel three variable loops (VL 1-3) form a bowl-like structure which is lined by positively charged residues, consituting the phosphoinositide-binding site. | ||
The PH domain of PDK1 possesses an additional extension (<scene name='56/568020/Bud/1'>'bud'</scene>) N-terminal to the standard PH domain fold (see Fig.1 blue section). This bud forms two additional β strands and one α helix and is an integral part of the overall fold. The two β strands β1' and β2'extend the β1 - β4 sheet in an antiparallel fashion through the formation of β sheet hydrogen bonds between β2'and β1. The α helix packs against this newly formed six stranded β sheet forming an additional <scene name='56/568020/Hydrophobic_core/1'>hydrophobic core</scene> outside of the standard PH domain fold. The bud binds to the β1 - β4 sheet by several additional hydrophobic contacts and buries more than 30% of the surface of the standard PH domain fold. <ref name="Structural" /> | The PH domain of PDK1 possesses an additional extension (<scene name='56/568020/Bud/1'>'bud'</scene>) N-terminal to the standard PH domain fold (see Fig.1 blue coloured section). This bud forms two additional β strands and one α helix and is an integral part of the overall fold. The two β strands β1' and β2'extend the β1 - β4 sheet in an antiparallel fashion through the formation of β sheet hydrogen bonds between β2'and β1. The α helix packs against this newly formed six stranded β sheet forming an additional <scene name='56/568020/Hydrophobic_core/1'>hydrophobic core</scene> outside of the standard PH domain fold. The bud binds to the β1 - β4 sheet by several additional hydrophobic contacts and buries more than 30% of the surface of the standard PH domain fold. <ref name="Structural" /> | ||
=== Binding Site === | === Binding Site === | ||
The <scene name='56/568020/Active_site/1'>binding site</scene> is formed by VL 1-3 which create a shallow, positively charged pocket at the open end of the β barrel. The structure was determined from a crystal grown in the presence of Ins(1,3,4,5)P<sub>4</sub>, which is the head group of PtdIns(3,4,5)P<sub>3</sub>. The phosphoinositide-binding site is lined by positively charged residues which contact the phosphates of Ins(1,3,4,5)P<sub>4</sub> by forming up to 11 hydrogen bonds. <scene name='56/568020/Arg472/1'>Arg472</scene> forms two hydrogen bonds to the phosphate in the D1 position (D1-phosphate). <scene name='56/568020/Argandlys/1'>Arg474 and Lys465</scene> contact the D3-phosphate with three hydrogen bonds in total (two for Arg474, one for Lys465). <scene name='56/568020/Lys495/1'>Lys495</scene> is also in close proximity (4.4 Å) of the D3-phosphate and is able to interact with it. <scene name='56/568020/Lys_tyr_and_arg/2'>Lys465, Tyr486 and Arg521</scene> form up to four hydrogen bonds to the D4-phosphate. The D5-phosphate is contacted by <scene name='56/568020/Lys467/1'>Lys467</scene> with one hydrogen bond. The D2- and D6-hydroxy groups show no direct interactions with the protein.<ref name="Structural" /> | The <scene name='56/568020/Active_site/1'>binding site</scene> is formed by VL 1-3 which create a shallow, positively charged pocket at the open end of the β barrel. The structure was determined from a crystal grown in the presence of Ins(1,3,4,5)P<sub>4</sub>, which is the head group of PtdIns(3,4,5)P<sub>3</sub>, a high affinity substrate of the PH domain of PDK1. The phosphoinositide-binding site is lined by positively charged residues which contact the phosphates of Ins(1,3,4,5)P<sub>4</sub> by forming up to 11 hydrogen bonds. <scene name='56/568020/Arg472/1'>Arg472</scene> forms two hydrogen bonds to the phosphate in the D1 position (D1-phosphate). <scene name='56/568020/Argandlys/1'>Arg474 and Lys465</scene> contact the D3-phosphate with three hydrogen bonds in total (two for Arg474, one for Lys465). <scene name='56/568020/Lys495/1'>Lys495</scene> is also in close proximity (4.4 Å) of the D3-phosphate and is able to interact with it. <scene name='56/568020/Lys_tyr_and_arg/2'>Lys465, Tyr486 and Arg521</scene> form up to four hydrogen bonds to the D4-phosphate. The D5-phosphate is contacted by <scene name='56/568020/Lys467/1'>Lys467</scene> with one hydrogen bond. The D2- and D6-hydroxy groups show no direct interactions with the protein.<ref name="Structural" /> | ||
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=== Interaction with Ins(1,3,4,5)P<sub>4</sub> === | === Interaction with Ins(1,3,4,5)P<sub>4</sub> === | ||
In the PDK1 PH domain Ins(1,3,4,5)P<sub>4</sub> complex Ins(1,3,4,5)P<sub>4</sub> interacts with protein side chains only. This results in a significantly reduced number of protein-ligand hydrogen bonds (a total of 11) compared to PH domain Ins(1,3,4,5)P<sub>4</sub> complexes of other proteins which form 15 to 16 hydrogen bonds. | |||
In the PDK1 Ins(1,3,4,5)P<sub>4</sub>-binding pocket a layer of five-ordered water molecules (B-factors) seperate Ins(1,3,4,5)P<sub>4</sub> from the protein (see Fig.2). The water molecules mediate a number of hydrogen bonds from Ins(1,3,4,5)P<sub>4</sub> to the protein. For example binding of the D2-hydroxyl group takes place via an ordered water molecule. But only one of these five water molecules is also conserved in the PH domains of other proteins contacting the D3-phosphate (see Fig.2, coloured yellow). | |||
