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Carbohydrate Esterase Family 2

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Carbohydrate Esterase Family CE2
Clan
Mechanism
Active site residues
CAZy DB link
http://www.cazy.org/CE3.html

Substrate specificities

All of the well characterized carbohydrate esterase family 2 enzymes have been shown to remove acetate groups from the synthetic molecule, 4-nitrophenyl acetate (4p-NP-AcC) (Montanier et al. 2009; Till et al. 2013). Contrary to typical acetyl xylan esterases, CE2 family members are shown to have a strong preference for the deacetylation of xylopyranosides at positions 3 and 4 instead of the typical deacetylation at position 2. CE2 family members were also shown to have significant preference for deacetylation of glucopyranosyl and mannopyranosyl residues at the 6-O position and significantly preferred deacetylation of glucopyranosyl and mannopyranosyl residues relative to the deacetylation of xylopyranosides. For these reasons CE2 family members are considered to be 6-de-O-acetylases (Topakas et al. 2010).

Catalytic Residues

Most CE2 family members contain a catalytic dyad (Ser-His) as opposed to a catalytic triad that is typically found in esterases. CjCE2A is an exception and contains a Ser-His-Asp catalytic triad (Montanier et al. 2009). The structurally characterized, CtCE2, CjCE2B, and Est2A are examples of CE2 enzymes that contain catalytic dyads of conserved serine and histidine residues and lack the aspartate residue found in the triad. Without the support of an aspartate residue, the histidine residue of the catalytic dyads are supported by main-chain carbonyl groups provided by a backbone amino acid. The catalytic aspartate residue that would commonly complete the catalytic triad simply does not exist in many CE2 members. When CE2 enzymes contain a potential catalytic aspartate residue, often enough, there also exists a tryptophan residue that sits between the catalytic histidine and aspartate residues thereby prevent the aspartate residue from completing the triad. CE2 enzymes also possess an aromatic residue (either a tyrosine or a tryptophan residue) above their binding clefts that promote greater substrate specificity (Montanier et al. 2009; Till et al. 2013). Lastly, the oxyanion hole is comprised of the catalytic serine residue, a glycine, and an asparagine residue that appears to be invariant across the CE2 family and that of other related acetyl-esterases (Montanier et al. 2009; Till et al. 2013).

Kinetics and Mechanism

Three-dimensional structures

There are four reported structures for the CE2 family. These structures are all reported to be α/β-hydrolases and include Clostridium thermocellum’s CtCE2 (PDB ID 2WAO), Cellvibrio japonicus’ CjCE2A (PDB ID 2WAA) and CjCE2B (PDB ID 2W9X), and Butyrivibrio proteoclasticus’ Est2A (PDB ID 3U37). They contain an N-terminal β-sheet “jelly-roll” domain that acts as a carbohydrate binding domain (CBM) and is linked to a C-terminal domain that contains a α/β-hydrolase fold (SGNH-hydrolase motif) (Montanier et al. 2009; Till et al. 2013). This α/β-hydrolase fold consists of a three layered α/β stack composed of five beta strands arranged in parallel that form a central β-sheet, which is packed between α-helicies. In the case of CtCE2, CjCE2A, and CjCE2B, the sheet has 5 α-helices in total packed on each side (Montanier et al. 2009), but Est2A, has 9 α-helices packing both sides of the β-sheet. The common structure of the N-terminal B-sheet “jelly-roll” domain across CE2 enzymes is comprised of two opposing β-sheets that have 4 and 5 β-strands, respectively (Till et al. 2013).


The CE2 family members are typically monomeric, but there are some exceptions. Specifically, Est2A has been found to form tetramers that combine to make an overall octameric structure (Montanier et al. 2009; Till et al. 2013). The overall structure of CtCE2 also displayed that this domain is connected to the C-terminal end of a GH5 family cellulase protein, CtCel5C (PDB ID 4IM4), which make up a modular protein, called, CtCel5C-CE2. This protein is incorporated into cell-wall degrading cellulosome in C. thermocellum (Montanier et al. 2009).

Family Firsts

First characterized
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First mechanistic insight
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First 3-D structure
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References

  1. Faik A (2010). Xylan biosynthesis: news from the grass. Plant Physiol. 2010;153(2):396-402. DOI:10.1104/pp.110.154237 | PubMed ID:20375115 [Faik2010]