Glycoside Hydrolase Family 58

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• Author: ^^^Warren Wakarchuk^^^
• Responsible Curator: ^^^Warren Wakarchuk^^^

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Glycoside Hydrolase Family GH58
Clan	none
Mechanism	inverting
Active site residues	not known
CAZy DB link
http://www.cazy.org/fam/GH58.html

Substrate specificities

Glycoside hydrolases of family 58 are endo-N-acetylneuraminidases (also termed endo-sialidases). The capsular coat surrounding E. coli K1, is poly-alpha-2,8-sialic acid and protects the bacterium from the host immune system, but it also acts as an anchor point for bacteriophage infection. A range of bacteriophages specific to this E. coli capsule type have been described which have tailspike enzymes that degrade the capuslar material [1, 2, 3]

Kinetics and Mechanism

Family 58 endo-sialidases are specific for polysialic acid and form a unique family of endo-acting sialidases, all others previously reported being exo-acting. The recently determined structure of an endo-sialidase derived from bacteriophage K1F (endoNF)[4] revealed the active site to lack a number of the residues that are conserved in other sialidases, implying a new, endo-sialidase-specific catalytic mechanism. Using synthetic trifluoromethylumbelliferyl oligosialoside substrates kinetic parameters for hydrolysis at a single cleavage site were determined. Measurement of k_cat/K_m at a series of pH values revealed a dependence on a single protonated group of pK_a 5. Direct ¹H-NMR analysis of the hydrolysis of trifluoromethylumbelliferyl sialotrioside revealed that endoNF undergoes catalysis with and inverting mechanism [5].

Catalytic Residues

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Three-dimensional structures

The structure for the K1F tailspike enzyme has been solved by Stummeyer et al. [4]. The protein structure is similar to other phage tailspike endo-hydrolases in that it is a an extended molecule that is trimeric. The first tailspike enzyme to be studied in detail was the endo-rhamnosidase from the Salmonella phage P22 [6], which is in Family GH90.

Family Firsts

First sterochemistry determination

The stereochemical outcome of this reaction was determined in the Withers laboratory at UBC using synthetic substrates with a trifluoromethyl-umbelliferol fluorophore [5]. This determination showed that unlike all other known sialidases, the endo-sialidase uses an inverting mechanism.

First catalytic nucleophile identification

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First general acid/base residue identification

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First 3-D structure

The first structure determination was performed with a truncated version ofbthe K1F protein [4].

References

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