Variation of Antigen 43 self-association modulates bacterial compacting within aggregates and biofilms

Vo J, Ortiz G, Totsika M, Lo A, Hancock S, Whitten A, Hor L, Peters K, Ageorges V, Caccia N, Desvaux M, Schembri M, Paxman J, Heras B, npj Biofilms and Microbiomes 8(1) (2022) DOI

SASDKP3 – Alpha domain of autotransporter adhesin Ag43a from Escherichia coli strain UTI89 / UPEC (alpha-Ag43_UTI)

Putative autotransporter

MW^I(0)	69	kDa
MW^expected	119	kDa
V^Porod	79	nm³

log I(s) 5.87×10^-2 5.87×10^-3 5.87×10^-4 5.87×10^-5

Putative autotransporter small angle scattering data

s, nm^-1

Log plot Log-Log plot

ln I(s)

ln 5.87×10^-2 R_g: 3.7 nm 0 (3.7 nm)^-2 s²

(sR_g)²I(s)/I(0)

1.104 0 √3 sR_g

p(r)	R_g: 3.7 nm 0 D_max: 13.5 nm

Data validation

Experimental data range

p(r) fit to data

Metric

Value

s_min D_max / π

0.6

N_Shannon

Worse

Better

Metric

Value

p_cormap

χ²

0.964
0.978

Worse

Better

Fits and models

log I(s)	s, nm^-1
+3 Δ/σ -3	s, nm^-1

X-ray synchrotron radiation scattering data from the Ag43a alpha domain from UTI89 in 25 mM HEPES 150 mM NaCl, pH 7.0 were collected on the SAXS/WAXS beam line of the Australian Synchrotron (Melbourne, Australia) using a 2D Photon counting Pilatus 1M-W pixel detector (s = 4π sin θ/λ, where 2θ is the scattering angle). Thirty five successive 1 second frames were collected at 1.2 mg/ml. The data were normalized to the intensity of the transmitted beam and radially averaged and the scattering of the solvent-blank was subtracted. The solution is a mixture of different oligomeric states, and a monomer-dimer mixture was fit to the data. The modelling involved taking the monomer and dimer from the crystal structure (PDB ID 7KO9), and generating the scattering profiles for each in Crysol. The relative proportions of each and a constant background correction were optimised by minimising χ2 using the Solver function in Excel. Approximately 77% of the protein was determined to be in the monomeric state and 23% in the dimeric state, with χ2 = 8.8. The model reproduces the features of the curve, but the small systematic deviations are likely due to the presence of small amounts of higher order oligomers.

Putative autotransporter (alpha-Ag43_UTI)
Mol. type		Protein
Organism		Escherichia coli (strain UTI89 / UPEC)
Olig. state		Dimer
Mon. MW		59.4 kDa
Sequence		FASTA