PDBsum entry 1k6a

Hydrolase

PDB id: 1k6a

Contents

Protein chain

302 a.a. *

Waters ×199

* Residue conservation analysis

References listed in PDB file

Key reference

Title

Substrate specificity and subsite mobility in t. Aurantiacus xylanase 10a.

Authors

L.Lo leggio, S.Kalogiannis, K.Eckert, S.C.Teixeira, M.K.Bhat, C.Andrei, R.W.Pickersgill, S.Larsen.

Ref.

FEBS Lett, 2001, 509, 303-308. [DOI no: 10.1016/S0014-5793(01)03177-5]

PubMed id

11741607

Abstract

The substrate specificity of Thermoascus aurantiacus xylanase 10A (TAX) has been investigated both biochemically and structurally. High resolution crystallographic analyses at 291 K and 100 K of TAX complexes with xylobiose show that the ligand is in its alpha anomeric conformation and provide a rationale for specificity on p-nitrophenyl glycosides at the -1 and -2 subsites. Trp 275, which is disordered in uncomplexed structures, is stabilised by its interaction with xylobiose. Two structural subsets in family 10 are identified, which differ by the presence or absence of a short helical stretch in the eighth betaalpha-loop of the TIM barrel, the loop bearing Trp 275. This structural difference is discussed in the context of Trp 275 mobility and xylanase function.

Figure 1.
Fig. 1. Xylobiose and glycerol binding in TAX. TAX residues are shown for the XBCRYO structure in red, magenta (tryptophan residues forming the aromatic cage at subsite −1) or orange (catalytic glutamates). Xylobiose and water 75 in the XBCRYO structure is shown in blue. The sugar moieties occupying subsites −2, −1 and +1 in XBRT and water 319 are shown in cyan. Glycerol and water 203 from the GLC structure are shown in green. Potential hydrogen bonds with TAX (distances shorter than 3.2 Å) are coloured according to the same scheme as for the ligands. The substrate binding groove is oriented so that subsite −2 is at the top and subsite +1 is at the bottom. In panel b a Sigmaa F[obs]−F[calc] map calculated prior to incorporation of xylobiose in the model (blue) is shown for XBCRYO contoured at 2σ around 1.5 Å from xylobiose. In white the conformation of xylobiose bound to the P. simplicissimum xylanase (PDB code 1B3W), is shown for comparison. In panel c a simulated annealing 2F[obs]−F[calc] omit map is shown for the xylose at subsite +1 in the XBRT model (cyan) contoured at 0.7σ. The complexes of P. simplicissimum xylanase with xylotetraose (PDB code 1B3Y) and xylopentaose (PDB code 1B3Z) are shown in white, and the complex of P. fluorescens xylanase 10A with xylopentaose (PDB code 1E5N) is shown in purple for comparison.

Figure 2.
Fig. 2. Alternative conformations for Glu 46, Trp 275 and Arg 276 in the form II crystal.

The above figures are reprinted by permission from the Federation of European Biochemical Societies: FEBS Lett (2001, 509, 303-308) copyright 2001.

Secondary reference #1

Title

Anisotropic refinement of the structure of thermoascus aurantiacus xylanase i.

Authors

S.Teixeira, L.Lo leggio, R.Pickersgill, C.Cardin.

Ref.

Acta Crystallogr D Biol Crystallogr, 2001, 57, 385-392. [DOI no: 10.1107/S0907444900019089]

PubMed id

11223515

Figure 3.
Figure 3 In the top-left corner a side view of the ( )[8] TIM-barrel fold of TAXI can be seen. A zoom on the active site shows the side chains of some selected residues (blue), illustrating the environment around the active-site glutamates (red). The alternate conformations of Trp275 are coloured in green. The top right corner shows a top view along the barrel.

Figure 5.
Figure 5 Progress of the refinement. The residual R factor is defined as R = |F[o]| - |F[c]| / |F[o]|.

The above figures are reproduced from the cited reference with permission from the IUCr

Secondary reference #2

Title

High resolution structure and sequence of t. Aurantiacus xylanase i: implications for the evolution of thermostability in family 10 xylanases and enzymes with (beta)alpha-Barrel architecture.

Authors

L.Lo leggio, S.Kalogiannis, M.K.Bhat, R.W.Pickersgill.

Ref.

Proteins, 1999, 36, 295-306. [DOI no: 10.1002/(SICI)1097-0134(19990815)36:3<295::AID-PROT4>3.3.CO;2-Y]

PubMed id

10409823

Figure 2.
Figure 2. Comparison of the structures of TAX, XYLA and XYNZ in a region containing a hydrophobic cavity in XYLA. A hydrophobic cavity present in XYLA (a) is filled by Phe 18 in TAX (b), and by Met 780, Phe 792, and Met 794 in XYNZ (c). The XYLA cavity is shown as a blue net. The figure was produced with QUANTA using the cavity definition of GRASP.

Figure 5.
Figure 5. Stereo representations of typical final electron density maps for the two crystal forms. Two SIGMAA weighted 2F[obs] - F[calc] electron density maps are shown at 1.2 contour level, showing typical final density for the Form I crystal at 1.92 Å resolution a) and for the Form II crystal at 1.14 Å resolution b). The same region of the protein is shown in both views.

The above figures are reproduced from the cited reference with permission from John Wiley & Sons, Inc.

Secondary reference #3

Title

Crystal structure at 1.8 a resolution and proposed amino acid sequence of a thermostable xylanase from thermoascus aurantiacus.

Authors

R.Natesh, P.Bhanumoorthy, P.J.Vithayathil, K.Sekar, S.Ramakumar, M.A.Viswamitra.

Ref.

J Mol Biol, 1999, 288, 999. [DOI no: 10.1006/jmbi.1999.2727]

PubMed id

10329194

Figure 2.
Figure 2. (a) Quality of the 2 F[o]−F[c] omit map (contoured at 1σ level) around the active site Glu237 after 50 cycles of positional refinement, omitting a stretch of residues around it from the refinement and electron density map calculation. (b) A representative electron density map showing the quality of the 2F[o]−F[c] electron density map contoured at 1σ level.

Figure 6.
Figure 6. Hydrogen bond network around the active site with some of the conserved water molecules in the active site labelled.

The above figures are reproduced from the cited reference with permission from Elsevier