The overall structure of XynA
The full-length XynA protein was expressed and purified with removal of
nucleic acids by anion exchange column QHP. XynA contains 408 residues
and the molecular weight of the tagged full-length protein was 49.8 kDa
with an isoelectric point of 6.64.
It was identified as one monomer in
the solution by gel filtration (Figure
1A) .
We found that xylanase 10A (SlXyn10A, PDB
code
1V0L) fromS.
lividans has the highest sequence similarity (30%) with XynA by using
Phyre2
server (Gloster et al., 2004; Kelley, Mezulis, Yates, Wass, &
Sternberg, 2015). We used this
structure as a search model to solve the crystal structure of XynA by
molecular replacement method.
The
determined 2.3 Å crystal structure of XynA (residues 22-393) with aR free of 0.245 (Table 1 ). The space
group is P4122 , and one molecule was present in the asymmetric
unit. In the refined model, residues Leu52, Ala53, Asp359, Ala360,
Asn361, Gly378, and Glu379 are missing due to disordered configuration.
The XynA structure is composed of
12 α-helices and 14 β-sheets, among which α2-α11 helices and β3-β10
sheets form a classical (α/β)8 TIM-barrel fold(Figure 1B) . Overall, the mug-shaped XynA is comprised of an
N-terminal
domain (residues 1-67), GH10 domain (residues 68-326) and C-terminal
domain (residues 327-408) (Figure 1C) .
The
body of “mug” is GH10 domain, which has 3 α-helices separated by loop
regions, named α2, α4, and α7 (Figure 1B) . These three
α-helices are not affecting the overall conformation of
classical
(α/β)8 TIM-barrel fold.