3RRX

Crystal Structure of Q683A mutant of Exo-1,3/1,4-beta-glucanase (ExoP) from Pseudoalteromonas sp. BB1

Summary for 3RRX

• Entry DOI: 10.2210/pdb3rrx/pdb
• Related: 3F95 3USZ 3UT0
• Descriptor: Exo-1,3/1,4-beta-glucanase, CALCIUM ION, SODIUM ION, ... (5 entities in total)
• Functional Keywords: (alpha/beta)8 barrel, (alpha/beta)6 sheet, hydrolase
• Biological source: Pseudoalteromonas sp.
• Total number of polymer chains: 1
• Total formula weight: 90751.73

Authors

Nakatani, Y.,Cutfield, S.M.,Cutfield, J.F. (deposition date: 2011-05-01, release date: 2011-12-21, Last modification date: 2024-10-16)

Primary citation

Nakatani, Y.,Cutfield, S.M.,Cowieson, N.P.,Cutfield, J.F.
Structure and activity of exo-1,3/1,4-beta-glucanase from marine bacterium Pseudoalteromonas sp. BB1 showing a novel C-terminal domain
Febs J., 279:464-478, 2012

PubMed Abstract: Following the discovery of an exo-1,3/1,4-β-glucanase (glycoside hydrolase family 3) from a seaweed-associated bacterium Pseudoalteromonas sp. BB1, the recombinant three-domain protein (ExoP) was crystallized and its structure solved to 2.3 Å resolution. The first two domains of ExoP, both of which contribute to the architecture of the active site, are similar to those of the two-domain barley homologue β-d-glucan exohydrolase (ExoI) with a distinctive Trp-Trp clamp at the +1 subsite, although ExoI displays broader specificity towards β-glycosidic linkages. Notably, excision of the third domain of ExoP results in an inactive enzyme. Domain 3 has a β-sandwich structure and was shown by CD to be more temperature stable than the native enzyme. It makes relatively few contacts to domain 1 and none at all to domain 2. Two of the domain 3 residues involved at the interface, Q683 (forming one hydrogen bond) and Q676 (forming two hydrogen bonds) were mutated to alanine. Variant Q676A retained about half the activity of native ExoP, but the Q683A variant was severely attenuated. The crystal structure of Q683A-ExoP indicated that domain 3 was highly mobile and that Q683 is critical to the stabilization of ExoP by domain 3. Small-angle X-ray scattering data lent support to this proposal. Domain 3 does not appear to be an obvious carbohydrate-binding domain and is related neither in sequence nor structure to the additional domains characterized in other glycoside hydrolase 3 subgroups. Its major role appears to be for protein stability but it may also help orient substrate.

PubMed: 22129429
DOI: 10.1111/j.1742-4658.2011.08439.x

Experimental method

X-RAY DIFFRACTION (1.9 Å)