Human MICAL1: activation by the small GTPase Rab8 and small-angle X-ray scattering studies on the oligomerization state of MICAL1 and its complex with Rab8.

Esposito A, Ventura V, Petoukhov MV, Rai A, Svergun DI, Vanoni MA, Protein Sci (2018) Europe PMC

SASDDT9 – NADPH oxidase (H2O2 producing and [F-actin] oxidizing) MICAL1 (monomer) in complex with Ras-related protein Rab 8 (MICAL1-Rab8 complex)

[F-actin]-monooxygenase MICAL1 (monomer)
Ras-related protein 8
MWexperimental 142 kDa
MWexpected 138 kDa
VPorod 234 nm3
log I(s) 8.78×103 8.78×102 8.78×101 8.78×100
[F-actin]-monooxygenase MICAL1 (monomer) Ras-related protein 8 small angle scattering data  s, nm-1
ln I(s)
[F-actin]-monooxygenase MICAL1 (monomer) Ras-related protein 8 Guinier plot ln 8.79×103 Rg: 3.7 nm 0 (3.7 nm)-2 s2
(sRg)2I(s)/I(0)
[F-actin]-monooxygenase MICAL1 (monomer) Ras-related protein 8 Kratky plot 1.104 0 √3 sRg
p(r)
[F-actin]-monooxygenase MICAL1 (monomer) Ras-related protein 8 pair distance distribution function Rg: 3.6 nm 0 Dmax: 11.8 nm

Data validation

Fits and models

log I(s)
 s, nm-1
[F-actin]-monooxygenase MICAL1 (monomer) Ras-related protein 8 CORAL model
log I(s)
 s, nm-1
[F-actin]-monooxygenase MICAL1 (monomer) Ras-related protein 8 CORAL model
log I(s)
 s, nm-1
[F-actin]-monooxygenase MICAL1 (monomer) Ras-related protein 8 CORAL model
Synchrotron SAXS data, I(s) vs s (where s = 4πsinθ/λ, and 2θ is the scattering angle), were measured from the MICAL1:Rab8 complex using continuous-flow size-exclusion chromatography SAXS (SEC-SAXS) on the EMBL-P12 beam line at the PETRA III storage ring (Hamburg, Germany). Data were collected a using a Pilatus 2M detector at a sample-detector distance of 3 m and at a wavelength of λ = 0.124 nm. The SEC mobile phase (0.5 ml/min) consisted of 20 mM Hepes/NaOH, pH 7.5, 50 mM NaCl, 2 mM MgCl2, 2 mM DTT, pH 7.5 (23°C). The sample load concentration onto the SEC column was 10 mg/ml (75 µl injection volume). The SAXS data measured from the SEC-elution (sample peak and buffer; 3600 successive 1 s frames) were normalized to the intensity of the transmitted beam and radially averaged. The scattering of an appropriate solvent-blank was subtracted from the sample frames to produce the scaled and averaged data displayed in this entry.

1:1 Complex between Mical1 and Rab8. Note: The protein in the SAXS experiments is Rab8, however the protein used for modelling is Rab10.

[F-actin]-monooxygenase MICAL1 (monomer) (Mical1)
Mol. type   Protein
Organism   Homo sapiens
Olig. state   Monomer
Mon. MW   117.9 kDa
 
UniProt   Q8TDZ2
Sequence   FASTA
 
Ras-related protein 8 (Rab8)
Mol. type   Protein
Olig. state   Monomer
Mon. MW   20.2 kDa
 
UniProt   P61026 (1-175)
Sequence   FASTA