Molecular basis of crosstalk in nuclear receptors: heterodimerization between PXR and CAR and the implication in gene regulation

Shirish chodankar.

SASDNZ3 – Heterodimerization between the nuclear receptor subfamily proteins PXR and CAR

Nuclear receptor subfamily 1 group I member 2
Nuclear receptor subfamily 1 group I member 3
MWexperimental 51 kDa
MWexpected 72 kDa
VPorod 56 nm3
log I(s) 3.17×100 3.17×10-1 3.17×10-2 3.17×10-3
Nuclear receptor subfamily 1 group I member 2 Nuclear receptor subfamily 1 group I member 3 small angle scattering data  s, nm-1
ln I(s)
Nuclear receptor subfamily 1 group I member 2 Nuclear receptor subfamily 1 group I member 3 Guinier plot ln 3.18×100 Rg: 2.7 nm 0 (2.7 nm)-2 s2
(sRg)2I(s)/I(0)
Nuclear receptor subfamily 1 group I member 2 Nuclear receptor subfamily 1 group I member 3 Kratky plot 1.104 0 3 sRg
p(r)
Nuclear receptor subfamily 1 group I member 2 Nuclear receptor subfamily 1 group I member 3 pair distance distribution function Rg: 2.7 nm 0 Dmax: 10.1 nm

Data validation


Fits and models


log I(s)
 s, nm-1
Nuclear receptor subfamily 1 group I member 2 Nuclear receptor subfamily 1 group I member 3 DAMMIF model
Nuclear receptor subfamily 1 group I member 2 Nuclear receptor subfamily 1 group I member 3 DAMFILT model

Synchrotron SAXS data from solutions of the complex formed between the nuclear receptor subfamily proteins PXR and CAR in 25 mM HEPES, 150 mM NaCl, 5% glycerol, 5 mM DTT, pH 7.9 were collected on the 16-ID (LiX) beam line at the National Synchrotron Light Source (NSLS-II; Brookhaven, NY, USA) using a Pilatus3 S 1M detector at a wavelength of λ = 0.081 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 65.00 μl sample was injected at a 0.50 ml/min flow rate onto a GE Superdex 200 Increase 5/150 column at 10°C. The data were normalized to the intensity of the transmitted beam and radially averaged; the scattering of the solvent-blank was subtracted. Data were recorded at two detector positions, SAXS and WAXS, and subsequently merged.

The PXR protein construct contains a tethered segment at the C-terminus. The CAR protein construct contains a tethered segment at the C-terminus and relates to isoform 5 (UniProt Q14994-5).

Nuclear receptor subfamily 1 group I member 2 (PXR)
Mol. type   Protein
Organism   Homo sapiens
Olig. state   Monomer
Mon. MW   39.1 kDa
 
UniProt   O75469 (130-434)
Sequence   FASTA
 
Nuclear receptor subfamily 1 group I member 3 (CAR)
Mol. type   Protein
Organism   Homo sapiens
Olig. state   Monomer
Mon. MW   32.5 kDa
 
UniProt   Q14994 (103-352)
Sequence   FASTA