Evolutionary and molecular basis of ADP-ribosylation reversal by zinc-dependent macrodomains

Ariza A, Liu Q, Cowieson N, Ahel I, Filippov D, Rack J, Journal of Biological Chemistry :107770 (2024) DOI

SASDTZ8 – Streptococcus pyogenes Protein-ADP-ribose hydrolase: glycine cleavage system H-like protein complex

Glycine cleavage system H-like protein
Protein-ADP-ribose hydrolase (D13G, Y23S, T61A, I114S, R177H, I246T)
MWexperimental 40 kDa
MWexpected 44 kDa
VPorod 53 nm3
log I(s) 7.44×10-3 7.44×10-4 7.44×10-5 7.44×10-6
Glycine cleavage system H-like protein Protein-ADP-ribose hydrolase (D13G, Y23S, T61A, I114S, R177H, I246T) small angle scattering data  s, nm-1
ln I(s)
Glycine cleavage system H-like protein Protein-ADP-ribose hydrolase (D13G, Y23S, T61A, I114S, R177H, I246T) Guinier plot ln 7.45×10-3 Rg: 2.4 nm 0 (2.4 nm)-2 s2
(sRg)2I(s)/I(0)
Glycine cleavage system H-like protein Protein-ADP-ribose hydrolase (D13G, Y23S, T61A, I114S, R177H, I246T) Kratky plot 1.104 0 √3 sRg
p(r)
Glycine cleavage system H-like protein Protein-ADP-ribose hydrolase (D13G, Y23S, T61A, I114S, R177H, I246T) pair distance distribution function Rg: 2.5 nm 0 Dmax: 8.4 nm

Data validation

Fits and models

log I(s)
 s, nm-1
Glycine cleavage system H-like protein Protein-ADP-ribose hydrolase (D13G, Y23S, T61A, I114S, R177H, I246T) ALPHAFOLD model
log I(s)
 s, nm-1
Glycine cleavage system H-like protein Protein-ADP-ribose hydrolase (D13G, Y23S, T61A, I114S, R177H, I246T) MONSA model
Synchrotron SAXS data from solutions of Streptococcus pyogenes Protein-ADP-ribose hydrolase: glycine cleavage system H-like protein complex in 50 mM Tris-HCl, 200 mM NaCl, 1 mM DTT, 5% (v/v) glycerol, pH 8 were collected on the B21 beam line at the Diamond Light Source (Didcot, UK) using a Pilatus 2M detector at a sample-detector distance of 4.0 m and 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 45.00 μl sample at 6 mg/ml was injected at a 0.16 ml/min flow rate onto a Shodex KW403-4F column at 15°C. 600 successive 3 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.

Glycine cleavage system H-like protein (SpyGcvH-L)
Mol. type   Protein
Organism   Streptococcus pyogenes serotype M5 (strain Manfredo)
Olig. state   Monomer
Mon. MW   13.2 kDa
 
UniProt   P0DN69 (1-110)
Sequence   FASTA
 
Protein-ADP-ribose hydrolase (D13G, Y23S, T61A, I114S, R177H, I246T) (SpyMacro)
Mol. type   Protein
Organism   Streptococcus pyogenes serotype M5 (strain Manfredo)
Olig. state   Monomer
Mon. MW   30.4 kDa
 
UniProt   P0DN70 (1-270)
Sequence   FASTA