Chaotic advection mixer for capturing transient states of diverse biological macromolecular systems with time-resolved small-angle X-ray scattering

Zielinski K, Katz A, Calvey G, Pabit S, Milano S, Aplin C, San Emeterio J, Cerione R, Pollack L, IUCrJ 10(3):363-375 (2023) DOI

SASDRB4 – GAC rRNA + L11 Protein: Time-Resolved 63 ms

58 nucleotide RNA L11-binding domain from E. coli 23S rRNA
50S ribosomal protein L11
MWexperimental 21 kDa
MWexpected 34 kDa
VPorod 33 nm3
log I(s) 1.76×101 1.76×100 1.76×10-1 1.76×10-2
58 nucleotide RNA L11-binding domain from E. coli 23S rRNA 50S ribosomal protein L11 small angle scattering data  s, nm-1
ln I(s)
58 nucleotide RNA L11-binding domain from E. coli 23S rRNA 50S ribosomal protein L11 Guinier plot ln 1.77×101 Rg: 2.4 nm 0 (2.4 nm)-2 s2
(sRg)2I(s)/I(0)
58 nucleotide RNA L11-binding domain from E. coli 23S rRNA 50S ribosomal protein L11 Kratky plot 1.104 0 √3 sRg
Dmax: 11.5 nm

Data validation

There are no models related to this curve.

Synchrotron SAXS data from solutions of GAC rRNA + L11 Protein: Time-Resolved 63 ms in 10 mM Na-MOPSO, 100 mM KCl, pH 6.5 were collected on the G1 beam line at the Cornell High Energy Synchrotron Source (CHESS) storage ring (Ithaca, NY, USA) using a Pilatus 100K detector at a wavelength of λ = 0.10972 nm (I(s) vs s, where s = 4πsinθ/λ, and 2θ is the scattering angle). Solute concentrations ranging between 2.3 and 3.1 mg/ml were measured at 20°C. 20 successive 5 second frames were collected. The data were normalized to the intensity of the transmitted beam and radially averaged; the scattering of the solvent-blank was subtracted.

GAC rRNA (60-80 micromolar) and L11 protein (80-100 micromolar) + 10 mM MgCl2 were mixed together with a Kenics-style, chaotic advection mixer. Molecular weight determined by Vc

58 nucleotide RNA L11-binding domain from E. coli 23S rRNA (GAC rRNA)
Mol. type   RNA
Organism   Escherichia coli
Olig. state   Monomer
Mon. MW   18.8 kDa
Sequence   FASTA
 
50S ribosomal protein L11 (L11 protein)
Mol. type   Protein
Organism   Thermus thermophilus
Olig. state   Monomer
Mon. MW   15.5 kDa
 
UniProt   P36238 (1-147)
Sequence   FASTA