1E5N

E246C mutant of P fluorescens subsp. cellulosa xylanase A in complex with xylopentaose

Summary for 1E5N

• Entry DOI: 10.2210/pdb1e5n/pdb
• Related: 1CLX 1QLD 1XYS
• Descriptor: ENDO-1,4-BETA-XYLANASE A, beta-D-xylopyranose-(1-4)-beta-D-xylopyranose-(1-4)-beta-D-xylopyranose-(1-4)-beta-D-xylopyranose-(1-4)-beta-D-xylopyranose, CALCIUM ION (3 entities in total)
• Functional Keywords: glycosyl hydrolase, family 10, xylan degradation, hydrolase
• Biological source: PSEUDOMONAS FLUORESCENS
• Total number of polymer chains: 2
• Total formula weight: 78534.55

Authors

Lo Leggio, L.,Jenkins, J.A.,Harris, G.W.,Pickersgill, R.W. (deposition date: 2000-07-27, release date: 2000-12-08, Last modification date: 2023-12-13)

Primary citation

Leggio, L.L.,Jenkins, J.,Harris, G.W.,Pickersgill, R.W.
X-ray crystallographic study of xylopentaose binding to Pseudomonas fluorescens xylanase A.
Proteins, 41:362-373, 2000

PubMed Abstract: The structure of the complex between a catalytically compromised family 10 xylanase and a xylopentaose substrate has been determined by X-ray crystallography and refined to 3.2 A resolution. The substrate binds at the C-terminal end of the eightfold betaalpha-barrel of Pseudomonas fluorescens subsp. cellulosa xylanase A and occupies substrate binding subsites -1 to +4. Crystal contacts are shown to prevent the expected mode of binding from subsite -2 to +3, because of steric hindrance to subsite -2. The loss of accessible surface at individual subsites on binding of xylopentaose parallels well previously reported experimental measurements of individual subsites binding energies, decreasing going from subsite +2 to +4. Nine conserved residues contribute to subsite -1, including three tryptophan residues forming an aromatic cage around the xylosyl residue at this subsite. One of these, Trp 313, is the single residue contributing most lost accessible surface to subsite -1, and goes from a highly mobile to a well-defined conformation on binding of the substrate. A comparison of xylanase A with C. fimi CEX around the +1 subsite suggests that a flatter and less polar surface is responsible for the better catalytic properties of CEX on aryl substrates. The view of catalysis that emerges from combining this with previously published work is the following: (1) xylan is recognized and bound by the xylanase as a left-handed threefold helix; (2) the xylosyl residue at subsite -1 is distorted and pulled down toward the catalytic residues, and the glycosidic bond is strained and broken to form the enzyme-substrate covalent intermediate; (3) the intermediate is attacked by an activated water molecule, following the classic retaining glycosyl hydrolase mechanism.

PubMed: 11025547

Experimental method

X-RAY DIFFRACTION (3.2 Å)