Crosstalk between regulatory elements in disordered TRPV4 N-terminus modulates lipid-dependent channel activity

Goretzki B, Wiedemann C, McCray B, Schäfer S, Jansen J, Tebbe F, Mitrovic S, Nöth J, Cabezudo A, Donohue J, Jeffries C, Steinchen W, Stengel F, Sumner C, Hummer G, Hellmich U, Nature Communications 14(1) (2023) DOI

SASDQJ8 – G. gallus TRPV4 N-terminal Domain, deletion mutant ∆N97

Transient receptor potential cation channel subfamily V member 4
MWexperimental 32 kDa
MWexpected 33 kDa
VPorod 41 nm3
log I(s) 1.82×103 1.82×102 1.82×101 1.82×100
Transient receptor potential cation channel subfamily V member 4 small angle scattering data  s, nm-1
ln I(s)
Transient receptor potential cation channel subfamily V member 4 Guinier plot ln 1.82×103 Rg: 2.5 nm 0 (2.5 nm)-2 s2
(sRg)2I(s)/I(0)
Transient receptor potential cation channel subfamily V member 4 Kratky plot 1.104 0 √3 sRg
p(r)
Transient receptor potential cation channel subfamily V member 4 pair distance distribution function Rg: 2.8 nm 0 Dmax: 11.9 nm

Data validation

Fits and models

log I(s)
 s, nm-1
G. gallus TRPV4 N-terminal Domain, deletion mutant ∆N97 Rg histogram Rg, nm
Transient receptor potential cation channel subfamily V member 4 PYMOL model
Transient receptor potential cation channel subfamily V member 4 PYMOL model
Transient receptor potential cation channel subfamily V member 4 PYMOL model
Transient receptor potential cation channel subfamily V member 4 EOM/RANCH model
Synchrotron SAXS data from solutions of G. gallus TRPV4 N-terminal Domain, deletion mutant ∆N97, in 20 mM Tris, 300 mM NaCl, 10 mM DTT, pH 7 were collected on the EMBL P12 beam line at PETRA III (DESY, Hamburg, Germany) using a Pilatus 6M detector at a sample-detector distance of 3 m and at a wavelength of λ = 0.124 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 40.00 μl sample at 6.5 mg/ml was injected at a 0.30 ml/min flow rate onto a GE Superdex 200 Increase 5/150 column at 20°C. 540 successive 0.995 second frames were collected through the entire SEC elution profile, where 22 frames were processed through the sample elution peak. The data were normalized to the intensity of the transmitted beam and radially averaged; the scattering of the solvent-blank was subtracted.

The molecule contains residues T98-D382 of G. gallus TRPV4, which includes a PIP2 binding site (K107-R111), a proline-rich region (T121-V134) and an Ankryin repeat domain. The individual, unsubtracted SEC-SAXS data frames and additional modelling results are made available in the full entry zip archive.

Transient receptor potential cation channel subfamily V member 4 (TRPV4)
Mol. type   Protein
Organism   Gallus gallus
Olig. state   Monomer
Mon. MW   32.5 kDa
 
UniProt   A0A1D5PXA5 (98-382)
Sequence   FASTA