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=='''Introduction'''== | =='''Introduction'''== | ||
Karyopherin Beta 2 (Kapβ2) is an importin that transports various cargo proteins into the nucleus through interactions with nucleoporins, which are proteins of the nuclear pore complex (NPC). One might overlook the significance of this protein but it actually plays a crucial role in the human body by mediating transport of RNA-binding proteins involved in [[Transcription and RNA Processing]], RNA transport and translation. The structure of Kapβ2 is composed of 20 antiparallel helices called HEAT repeats. These HEAT repeats contribute to Kapβ2’s large superhelical shape. The protein is shown to form two arches: one at the N-terminal and the other at the C-terminal. Through recognition of a nuclear localization signal (NLS) located on its cargo, Kapβ2 binds to its cargo via its C-terminal arch. Release of the cargo is mediated by RanGTP, which once bound, leads to a large movement of the 62-residue heat repeat 8 loop into the C-terminal arch. This conformational change results in the dissociation of the cargo as the b2 loop binds the NLS binding site. | Karyopherin Beta 2 (Kapβ2) is an importin that transports various cargo proteins into the nucleus through interactions with nucleoporins, which are proteins of the nuclear pore complex (NPC). One might overlook the significance of this protein but it actually plays a crucial role in the human body by mediating transport of RNA-binding proteins involved in [[Transcription and RNA Processing]], RNA transport and translation. The structure of Kapβ2 is composed of 20 antiparallel helices called HEAT repeats. These HEAT repeats contribute to Kapβ2’s large superhelical shape. The protein is shown to form two arches: one at the N-terminal and the other at the C-terminal. Through recognition of a nuclear localization signal (NLS) located on its cargo, Kapβ2 binds to its cargo via its C-terminal arch. Release of the cargo is mediated by RanGTP, which once bound, leads to a large movement of the 62-residue heat repeat 8 loop into the C-terminal arch. This conformational change results in the dissociation of the cargo as the b2 loop binds the NLS binding site. | ||
<StructureSection load= size=475 side='right' scene='37/372725/Kapb2/4'> | |||
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<font size=3> '''Structure of Kapβ2''' </font> | <font size=3> '''Structure of Kapβ2''' </font> | ||
Kapβ2 is a superhelix comprised of 20 HEAT repeats (the name HEAT derives from Huntington, Elongation factor 3 A subunit of protein phosphatase 2A and Tor1 kinase), each of which consists of <scene name='37/372725/Kapb2_with_nls_heat_repeat1/1'>two anti-parallel helices</scene> (shown in red). The electrostatic potential of the internal surface of Kapβ2 superhelix at the C-terminal arch is negative. <font color=' | Kapβ2 is a superhelix comprised of 20 HEAT repeats (the name HEAT derives from Huntington, Elongation factor 3 A subunit of protein phosphatase 2A and Tor1 kinase), each of which consists of <scene name='37/372725/Kapb2_with_nls_heat_repeat1/1'>two anti-parallel helices</scene> (shown in red). The electrostatic potential of the internal surface of Kapβ2 superhelix at the C-terminal arch is negative. <font color='plum'><b> HEAT repeats </b></font> <scene name='37/372725/Kapb2_hr_9-13/2'>9-13</scene> and <scene name='37/372725/Kapb2_hr_14-18/1'>14-18</scene> form the binding site of Kapβ2 cargos while repeats 1-8 constitute the Ran GTPase binding site. Ran GTPase, a small 216-residue protein, is found more frequently in the nucleus and enables cargos to be released from Kapβ2. | ||
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The PY-NLS of 2H4M contains a hydrophobic rather than a basic N-terminal motif. Hydrophobic interactions at the N-terminal motif of the PY-NLS include: | The PY-NLS of 2H4M contains a hydrophobic rather than a basic N-terminal motif. Hydrophobic interactions at the N-terminal motif of the PY-NLS include: | ||
<scene name='37/372725/Kapb2_hydrophobic_interactions/ | <scene name='37/372725/Kapb2_hydrophobic_interactions/4'>Phe273, Gly274, Pro275 and Met276 of the NLS with Trp730 and Ile773 of KapB2</scene>. Interactions of the C-terminal RX2-5PY motif of the NLS include: <scene name='37/372725/Kapb2_hydrophobic_interactions/6'>Arg284 of the NLS with Glu509 and Asp543 of KapB2.</scene>; <scene name='37/372725/Kapb2_hydrophobic_interactions/9'>Pro288 and Tyr289 of the NLS with Ala380, Ala381, Asp384, Leu419, Ile457, Trp460 and Arg464 of KapB2.</scene> | ||
Upon binding Kapβ2,the NLS gains structure, conforms to and makes contact with the internal surface of the KapB2 C-terminal arch. | Upon binding Kapβ2,the NLS gains structure, conforms to and makes contact with the internal surface of the KapB2 C-terminal arch. | ||
<font size=3> '''How does Kapβ2 pass through the NPC?''' </font> | |||
Once the cargo binds to Kapβ2, the complex travels through the NPC by interactions with the FG-Nups. | |||
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</StructureSection> | |||
<StructureSection load='1qbk' size='500' side='left' caption='KapB2 with Ran (PDB entry [[1qbk]])' scene='37/372725/1qbk/1'> | |||
=='''How does Kapβ2 release its cargo? '''== | |||
Ran is a GTP binding protein that is found in greater concentrations in the nucleus than in the cytoplasm. In the nucleus Ran Guanine Nucleotide Exchange Factor (ranGEF) is a protein that catalyzes the dissociation of GDP from Ran and subsequent phosphorylation. Ran-GTP undergoes conformational changes that allow it to have a greater affinity for Kapβ2 than Ran-GDP. These conformational changes occur in Ran between residues 30-47 and residues 65-80. | |||
RanGTP has an overall positive charge allowing it to bind to the highly negative core of the N-terminal arch Kapβ2. There are two distinct regions of polar interaction between KapB2 and RanGTP. These regions of Kapβ2 are defined as the N Interface found within HR1-3, and the Central Interface found within HR6, HR7, 62-residue loop of HR8, HR13 and HR14. | |||
Polar interactions at the N Interface include Ser22, Arg31, Glu161, Asp 164 and Ser165 of KapB2, and Trp64, Val45, Arg110, and Arg106 of RanGTP. | |||
Polar interactions at the Central Interface include Asn271, Glu275, Glu278, Glu332, Glu333, Asp334, Arg336, His340, Glu363, Ile364, Asp366, Asp367, Ile370, Ser371, Trp373, Lys377, Asp639, Glu682 of KapB2 and Asn143, Lys141, Arg140, Asn154, Asn156, Lys159, Asp148, Tyr155, Lys127, Lys134, Lys132, Arg129, His139, Tyr 146, Gln145, Asn143, Arg140, and Lys127 of RanGTP. | |||
When RanGTP binds to Kapβ2 conformational changes occur within the N-terminal arch and within a long (62 residue) loop of HEAT repeat 8. It is the changes in the 62-residue loop (residues 311-373) that is important in the dissociation of the NLS. It is thought that the 62- residue loop binds into the NLS binding site thus releasing the NLS. | |||
</StructureSection> | |||