Structural insights into the substrate-binding proteins Mce1A and Mce4A from Mycobacterium tuberculosis

Asthana P, Singh D, Pedersen J, Hynönen M, Sulu R, Murthy A, Laitaoja M, Jänis J, Riley L, Venkatesan R, IUCrJ 8(5) (2021) DOI

SASDK32 – Mammalian cell entry protein 1A (Mce1A38-454)

Mce-family protein Mce1A
n-Dodecyl-β-D-Maltopyranoside
MWexperimental 159 kDa
MWexpected 161 kDa
VPorod 362 nm3
log I(s) 1.47×10-1 1.47×10-2 1.47×10-3 1.47×10-4
Mce-family protein Mce1A n-Dodecyl-β-D-Maltopyranoside small angle scattering data  s, nm-1
ln I(s)
Mce-family protein Mce1A n-Dodecyl-β-D-Maltopyranoside Guinier plot ln 1.47×10-1 Rg: 5.3 nm 0 (5.3 nm)-2 s2
(sRg)2I(s)/I(0)
Mce-family protein Mce1A n-Dodecyl-β-D-Maltopyranoside Kratky plot 1.104 0 3 sRg
Dmax: 21.6 nm

Data validation


Fits and models


log I(s)
 s, nm-1
Mce-family protein Mce1A n-Dodecyl-β-D-Maltopyranoside CUSTOM IN-HOUSE model

log I(s)
 s, nm-1
Mce-family protein Mce1A n-Dodecyl-β-D-Maltopyranoside CUSTOM IN-HOUSE model

Synchrotron SAXS data from solutions of Mammalian cell entry protein 1A (Mce1A38-454) in 50 mM Tris, 500 mM NaCl, 10% Glycerol, 5mM DDM, 1 mM β-ME, pH 8 were collected on the B21 beam line at the Diamond Light Source storage ring (Didcot, UK) using a Pilatus 2M detector at a wavelength of λ = 0.1 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 55.00 μl sample at 5 mg/ml was injected at a 0.08 ml/min flow rate onto a GE Superdex 200 Increase 3.2/300 column . The data were normalized to the intensity of the transmitted beam and radially averaged; the scattering of the solvent-blank was subtracted.

Synchrotron SAXS data of the Mce+Helical+Tail domain of Mammalian cell entry protein (Mce) 1A collected at DLS beamline B21 using SEC-SAXS. The molecular mass (MW) of the protein-detergent complex was determined using Size exclusion chromatography-multi angle light scattering (SEC-MALS) coupled to refractive index measurements. The model files contain the protein-detergent molecules. The protein models generated using iTasser were fitted against the SAXS data using the in-house program, also taking in account the micelle contribution. The aggregation number for the extended model (220) is provided with the sample data and the aggregation number for the coiled-coil model is 200.

Mce-family protein Mce1A (Mce1A38_454)
Mol. type   Protein
Organism   Mycobacterium tuberculosis
Olig. state   Monomer
Mon. MW   48.2 kDa
 
UniProt   Q79FZ9 (38-454)
Sequence   FASTA
 
n-Dodecyl-β-D-Maltopyranoside (DDM)
Mol. type   Other
Olig. state   0
Mon. MW   0.5 kDa
Chemical formula