Sandbox Reserved 973: Difference between revisions
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<Structure load='4F3L' size='350' frame='true' align='right' caption='Insert caption here' scene='Insert optional scene name here' />' | <Structure load='4F3L' size='350' frame='true' align='right' caption='Insert caption here' scene='Insert optional scene name here' />' | ||
== Function and overall structure == | == Function and overall structure == | ||
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Basic helix-loop-helix domains of CLOCK and BMAL1 are especially composed by 2 C-terminal helices called α1 and α2 that are involved in the formation of a canonical four-helical bHLH bundle. This bond between the HLH domains helps to stabilize the heterodimeric complex as the core of the the bundle is very hydrophobic. The spatial arrangement of this assembly has a major role in the the E-box recognition. The α1 helices are responseable for the DNA binding and the aminoacids sequence is crucial. Site-directed mutagenesis experiments showed that some hydrophobic residues, leucine in particular, were necessary in ordre to interact with the major grive of DNA duplex.In fact, when Leu57 and Leu74 of CLOCK, and Leu95 and Leu115 of BMAL1 are mutated to glutamate, mutants show no transactivation activity anymore bécasse the ability to form stable four-helice bundle is reduced as we can observe it through a bimolecular fluorescence complementation (BiFC) assay. in addition most of these mutations tend to unsettle the full length hétérodimeric complex. | Basic helix-loop-helix domains of CLOCK and BMAL1 are especially composed by 2 C-terminal helices called α1 and α2 that are involved in the formation of a canonical four-helical bHLH bundle. This bond between the HLH domains helps to stabilize the heterodimeric complex as the core of the the bundle is very hydrophobic. The spatial arrangement of this assembly has a major role in the the E-box recognition. The α1 helices are responseable for the DNA binding and the aminoacids sequence is crucial. Site-directed mutagenesis experiments showed that some hydrophobic residues, leucine in particular, were necessary in ordre to interact with the major grive of DNA duplex.In fact, when Leu57 and Leu74 of CLOCK, and Leu95 and Leu115 of BMAL1 are mutated to glutamate, mutants show no transactivation activity anymore bécasse the ability to form stable four-helice bundle is reduced as we can observe it through a bimolecular fluorescence complementation (BiFC) assay. in addition most of these mutations tend to unsettle the full length hétérodimeric complex. | ||
=== PAS-A domain == | === PAS-A domain === | ||
PAS-A domains don't have the same conformation in the two subunits. In BMAL1, we can observe 3 loops involving about 60 residues whereas in CLOCK there are only 25 residues in a single loop. Nevertheless, this two PAS-A domains adopt a typical PAS fold. The core of these domains contains a five-stranded antiparallel β sheet (AβBβGβHβIβ) as well as numerous α helices (Cα, DαEαFα). They also contain an N-terminal A'α helix that does not belong to the canonical PAS fold. Those helices pack in between the β-sheet faces and are involved in the dimerization interactions. | PAS-A domains don't have the same conformation in the two subunits. In BMAL1, we can observe 3 loops involving about 60 residues whereas in CLOCK there are only 25 residues in a single loop. Nevertheless, this two PAS-A domains adopt a typical PAS fold. The core of these domains contains a five-stranded antiparallel β sheet (AβBβGβHβIβ) as well as numerous α helices (Cα, DαEαFα). They also contain an N-terminal A'α helix that does not belong to the canonical PAS fold. Those helices pack in between the β-sheet faces and are involved in the dimerization interactions. | ||
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== Effects on the circadian cycle== | == Effects on the circadian cycle== | ||
== Diseases == | == Diseases == | ||
== Structural highlights == | == Structural highlights == | ||
