Small-angle neutron scattering studies suggest the mechanism of BinAB protein internalization

Sharma M, Aswal V, Kumar V, Chidambaram R, IUCrJ 7(2) (2020) DOI

SASDF97 – Complex of Binary toxin receptor (Cqm1) with deuterated BinB component protein in 100% D2O

Binary toxin receptor Cqm1 protein
Deuterated BinB component of mosquito-larvicidal Binary toxin
MWI(0) 67 kDa
MWexpected 121 kDa
log I(s) 4.80×10-2 4.80×10-3 4.80×10-4 4.80×10-5
Binary toxin receptor Cqm1 protein Deuterated BinB component of mosquito-larvicidal Binary toxin small angle scattering data  s, nm-1
ln I(s)
Binary toxin receptor Cqm1 protein Deuterated BinB component of mosquito-larvicidal Binary toxin Guinier plot ln 4.80×10-2 Rg: 1.9 nm 0 (1.9 nm)-2 s2
(sRg)2I(s)/I(0)
Binary toxin receptor Cqm1 protein Deuterated BinB component of mosquito-larvicidal Binary toxin Kratky plot 1.104 0 3 sRg
p(r)
Binary toxin receptor Cqm1 protein Deuterated BinB component of mosquito-larvicidal Binary toxin pair distance distribution function Rg: 2.2 nm 0 Dmax: 7.3 nm

Data validation


Fits and models


log I(s)
 s, nm-1
Binary toxin receptor Cqm1 protein Deuterated BinB component of mosquito-larvicidal Binary toxin PDB model

log I(s)
 s, nm-1
Binary toxin receptor Cqm1 protein Deuterated BinB component of mosquito-larvicidal Binary toxin DAMMIN model

SANS data from solutions of Complex of Binary toxin receptor (Cqm1) with deuterated BinB component protein in 100% D2O in 25 mM HEPES, pH 7.5, 25 mM NaCl, in 100% D2O, pH 7.5 were collected using a Ingenuously built detector at a sample-detector distance of 2 m and at a wavelength of λ = 0.52 nm (I(s) vs s, where s = 4πsinθ/λ, and 2θ is the scattering angle). One solute concentration of 1.00 mg/ml was measured at 25°C. One 43200 second frame was collected. The data were normalized to the intensity of the transmitted beam and radially averaged; the scattering of the solvent-blank was subtracted.

This data is for a stable complex of receptor Cqm1 protein and deuterated BinB component of mosquito-larvicidal Binary (BinAB) toxin. Deuterated BinB (dBinB) protein was synthesized using E. coli based protein expression system grown in M9+ medium in D2O after three step adaption. The extent of deuteration for dBinB was estimated from contrast (p − s)2 values to be ~77%. Theoretical molecular weight of the dBinB protein includes 77% of D atoms. SANS analysis (DAMMIN and CRYSON fit) suggest that deuterated BinB (dBinB), expectedly, is fully matched out against the used solvent (100% D2O) and only monomer of Cqm1 is modelled from SANS experiments

Binary toxin receptor Cqm1 protein (Cqm1)
Mol. type   Protein
Organism   synthetic construct
Olig. state   Monomer
Mon. MW   64.7 kDa
Sequence   FASTA
 
Deuterated BinB component of mosquito-larvicidal Binary toxin (dBinB)
Mol. type   Protein
Organism   synthetic construct
Olig. state   Monomer
Mon. MW   56.4 kDa
Sequence   FASTA
 
PDB code   6K5P