Overall structure of fully assembled cyanobacterial KaiABC circadian clock complex by an integrated experimental-computational approach.

Yunoki Y, Matsumoto A, Morishima K, Martel A, Porcar L, Sato N, Yogo R, Tominaga T, Inoue R, Yagi-Utsumi M, Okuda A, Shimizu M, Urade R, Terauchi K, Kono H, Yagi H, Kato K, Sugiyama M, Commun Biol 5(1):184 (2022) Europe PMC

SASDNJ2 – Circadian clock KaiA-KaiB-KaiC ternary protein complex measured by SEC-SAXS

Circadian clock protein KaiA
Circadian clock protein KaiB
Circadian clock protein kinase KaiC (S431D mutant)
MWexperimental 810 kDa
MWexpected 821 kDa
VPorod 1650 nm3
log I(s) 4.48×10-1 4.48×10-2 4.48×10-3 4.48×10-4
Circadian clock protein KaiA Circadian clock protein KaiB Circadian clock protein kinase KaiC (S431D mutant) small angle scattering data  s, nm-1
ln I(s)
Circadian clock protein KaiA Circadian clock protein KaiB Circadian clock protein kinase KaiC (S431D mutant) Guinier plot ln 4.49×10-1 Rg: 7.0 nm 0 (7.0 nm)-2 s2
(sRg)2I(s)/I(0)
Circadian clock protein KaiA Circadian clock protein KaiB Circadian clock protein kinase KaiC (S431D mutant) Kratky plot 1.104 0 √3 sRg
p(r)
Circadian clock protein KaiA Circadian clock protein KaiB Circadian clock protein kinase KaiC (S431D mutant) pair distance distribution function Rg: 7.0 nm 0 Dmax: 24.5 nm

Data validation

Fits and models

log I(s)
 s, nm-1
Circadian clock protein KaiA Circadian clock protein KaiB Circadian clock protein kinase KaiC (S431D mutant) OTHER model
log I(s)
 s, nm-1
Circadian clock protein KaiA Circadian clock protein KaiB Circadian clock protein kinase KaiC (S431D mutant) OTHER model
Synchrotron SAXS data from solutions of the circadian clock KaiA-KaiB-KaiC ternary protein complex in 50 mM sodium phosphate buffer, 150 mM NaCl, 5 mM MgCl2, 0.5 mM EDTA, 1 mM ATP, 1 mM DTT, 50 mM arginine, 50 mM glutamine, pH 7.8 were collected on the BL-10C beam line at the Photon Factory (PF), High Energy Accelerator Research Organization (KEK, Tsukuba, Japan) using a Pilatus3 2M detector at a sample-detector distance of 3.0 m and at a wavelength of λ = 0.15 nm (I(s) vs s, where s = 4πsinθ/λ, and 2θ is the scattering angle). In-line size-exclusion chromatography (SEC) SAS was employed. The SEC parameters were as follows: A sample was injected onto a column at 25°C. The data were normalized to the intensity of the transmitted beam and radially averaged; the scattering of the solvent-blank was subtracted.

The molecular weight was determined by analytical ultracentrifugation (AUC). SEC column and SEC parameters: UNKNOWN. X-ray exposure time: UNKNOWN.

Circadian clock protein KaiA
Mol. type   Protein
Organism   Synechococcus elongatus (strain PCC 7942 / FACHB-805)
Olig. state   Dodecamer
Mon. MW   32.8 kDa
 
UniProt   Q79PF6 (1-284)
Sequence   FASTA
 
Circadian clock protein KaiB
Mol. type   Protein
Organism   Synechococcus elongatus (strain PCC 7942 / FACHB-805)
Olig. state   Hexamer
Mon. MW   11.8 kDa
 
UniProt   Q79PF5 (1-102)
Sequence   FASTA
 
Circadian clock protein kinase KaiC (S431D mutant)
Mol. type   Protein
Organism   Synechococcus elongatus (strain PCC 7942 / FACHB-805)
Olig. state   Hexamer
Mon. MW   59.5 kDa
 
UniProt   Q79PF4 (1-519)
Sequence   FASTA