PDBsum entry 6u4z

Hydrolase

PDB id: 6u4z

Contents

Protein chain

473 a.a.

Ligands

P6G

EDO

Waters ×370

PDB id:

6u4z

Name:

Hydrolase

Title:

Crystal structure of a family 76 glycoside hydrolase from a bovine bacteroides thetaiotaomicron strain

Structure:

Alpha-1,6-mannanase. Chain: a. Synonym: cellobiose 2-epimerase. Engineered: yes

Source:

Bacteroides thetaiotaomicron. Organism_taxid: 818. Gene: btheta7330_05006, dw011_13140. Expressed in: escherichia coli bl21. Expression_system_taxid: 511693

Resolution:

1.40Å R-factor: 0.181 R-free: 0.195

Authors:

D.R.Jones,D.W.Abbott

Key ref:

D.R.Jones et al. (2020). Analysis of Active Site Architecture and Reaction Product Linkage Chemistry Reveals a Conserved Cleavage Substrate for an Endo-alpha-mannanase within Diverse Yeast Mannans. J Mol Biol, 432, 1083-1097. PubMed id: 31945375 DOI: 10.1016/j.jmb.2019.12.048

Date:

26-Aug-19 Release date: 29-Apr-20

Protein chain

A0A0P0FW82  (A0A0P0FW82_BACT4) -  Alpha-1,6-mannanase from Bacteroides thetaiotaomicron

Seq: 514 a.a.

Struc: 473 a.a.*

* PDB and UniProt seqs differ at 1 residue position (black cross)

Enzyme reactions

• Enzyme class: E.C.5.1.3.11 - cellobiose epimerase.
• Reaction: D-cellobiose = beta-D-glucosyl-(1->4)-D-mannopyranose
• Cofactor: NAD

Molecule diagrams generated from .mol files obtained from the KEGG ftp site

Added reference

DOI no: 10.1016/j.jmb.2019.12.048

J Mol Biol 432:1083-1097 (2020)

PubMed id: 31945375

Analysis of Active Site Architecture and Reaction Product Linkage Chemistry Reveals a Conserved Cleavage Substrate for an Endo-alpha-mannanase within Diverse Yeast Mannans.

D.R.Jones, X.Xing, J.P.Tingley, L.Klassen, M.L.King, T.W.Alexander, D.W.Abbott.

ABSTRACT

Yeast α-mannan (YM) is a densely branched N-linked glycan that decorates the surface of yeast cell walls. Owing to the high degree of branching, cleavage of the backbone of YM appears to rely on the coupled action of side-chain-cleaving enzymes. Upon examining the genome sequences of bovine-adapted Bacteroides thetaiotaomicron strains, isolated for their ability to degrade YM, we have identified a tandem pair of genes inserted into an orphan pathway predicted to be involved in YM metabolism. Here, we investigated the activity of one of these enzymes, a predicted endo-mannanase from glycoside hydrolase (GH) family 76 (BtGH76-MD40). Purified recombinant BtGH76-MD40 displayed activity on structurally distinct YMs from Saccharomyces cerevisiae and Schizosaccharomyces pombe. Linkage analysis of released oligosaccharide products from S. cerevisiae and S. pombe mannan determined BtGH76-MD40 targets a specific linkage that is conserved in structurally diverse YM substrates. In addition, using two differential derivatization methods, we have shown that there is an absolute requirement for undecorated d-mannopyranose in the -1 subsite. Determination of the BtGH76-MD40 X-ray crystal structure and structural superimposition and molecular docking of a branched alpha-mannopentatose substrate supported these findings. In contrast, BtGH76-MD40 can accommodate extended side chains in the +1 and -2 subsites, highlighting that a single alpha-1,6-mannosyl residue is a prerequisite for activity, and cleavage occurs at the reducing end of the undecorated monosaccharide. Collectively these results demonstrate how acquisition of new enzymes within extant pathways contributes to the functional abilities of saccharolytic bacteria persisting in complex digestive ecosystems.