Structural Analysis of the Partially Disordered Protein EspK from Mycobacterium Tuberculosis

Gijsbers A, Sánchez-Puig N, Gao Y, Peters P, Ravelli R, Siliqi D, Crystals 11(1):18 (2020) DOI

SASDKR4 – EspK secretion protein - C-terminal domain

ESX-1 secretion-associated protein EspK
MWexperimental 28 kDa
MWexpected 30 kDa
VPorod 52 nm3
log I(s) 8.25×10-2 8.25×10-3 8.25×10-4 8.25×10-5
ESX-1 secretion-associated protein EspK small angle scattering data  s, nm-1
ln I(s)
ESX-1 secretion-associated protein EspK Guinier plot ln 8.26×10-2 Rg: 2.2 nm 0 (2.2 nm)-2 s2
(sRg)2I(s)/I(0)
ESX-1 secretion-associated protein EspK Kratky plot 1.104 0 √3 sRg
p(r)
ESX-1 secretion-associated protein EspK pair distance distribution function Rg: 2.2 nm 0 Dmax: 8.4 nm

Data validation

Fits and models

log I(s)
 s, nm-1
ESX-1 secretion-associated protein EspK GASBOR model
ESX-1 secretion-associated protein EspK ITASSER model
The structural characterisation of C-terminal region of EspK was performed by SAXS coupled to an online size exclusion chromatography equilibrated with 20 mM Tris-HCl pH 8.0, 300 mM NaCl. The data were collected in the bioSAXS beamline B21 at Diamond Light Source, Harwell, United Kingdom. Protein sample consisting of 50 µL at a concentration of 13 mg mL-1 was injected onto a Shodex KW-403 size exclusion column attached to an Agilent HPLC at a flow rate of 0.08 mL min-1. The eluted protein was directed through a 1.6 mm diameter quartz capillary cell held in vacuum. Data acquisition consisted of 580 frames (with 3 s exposure time) using a PILATUS 2M detector at a distance of 4.014 m from the sample. Collected two-dimensional images were corrected for variations in beam current, normalized for time exposure, and processed into one-dimensional scattering curves using GDA and the DAWN software (Diamond Light Source, UK). Background was manually subtracted using the program ScÅtter (http://www.bioisis.net/scatter). SevERAL ab-inito models were obtained by I-TASSER and next were compared with a low resolution model obtained by GASBOR program (ATSAS)

ESX-1 secretion-associated protein EspK (Espk_CT)
Mol. type   Protein
Organism   Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv)
Olig. state   Monomer
Mon. MW   30.3 kDa
 
UniProt   P9WJC1 (517-729)
Sequence   FASTA