Chitosanase families


The family GH75 includes essentially enzymes of fungal origin, with some members originating from actinobacteria. While chitosanase activity of fungal species has been reported earlier by Monaghan et al. (87) and Fenton and Eveleigh (25), the first molecular sequence belonging to GH75 family was published in 1996 by Shimosaka et al (21). The producing microorganism, Fusarium solani is now called Nectria haematococca var. brevicona. The same research group determined the sequence of a chitosanase from Aspergillus oryzae (51). These two chitosanases are also similar to the enzyme from Aspergillus fumigatus (52,107), and to the chitosanase from the entomopathogenic fungus Metarhizium anisopliae (59). Because these chitosanases had no detectable sequence homology with other chitosanase families, they were classified in a different glycoside hydrolase family: the family GH75.

In 2002 and 2003, the publication of complete genome sequences of the actinomycetes Streptomyces coelicolor A3(2) and Streptomyces avermitilis (108, 109) revealed that S. avermitilis has two genes (SAV_1288 and SAV_1850) while S. coelicolor A3(2) has one gene (SCO7070) encoding chitosanases of GH family 75. More recently, the chitosanase encoded by the SAV_1850 chitosanase (also named SaCsn75A or Csn75A) have been expressed in recombinant E. coli and substrate-binding preference of this enzyme has been investigated (122) (see also the "Cleavage specificity" section).

Currently, GH75 family includes some sixty enzymes of fungal or bacterial origin. The enzymes which were characterized by biochemical techniques showed exclusively a chitosanase activity. The alignment of 10 GH75 protein sequences revealed two highly conserved clusters of amino acid residues: D[VI]DCDG and GEAS (see also (107)). The same aligment showed that GH75 enzymes from fungal and actinomycetal origin are grouped in two well separated clusters, as shown on the unrooted phylogenetic tree below:

Phylogenetic unrooted tree calculated from a Clustal Omega (123) alignment of 10 GH75 protein sequences.
SAV1288 and SAV1850, genes from Streptomyces avermitilis; SGR_1238, gene from Streptomyces griseus subsp. griseus NBRC 13350; SCAB_83781, gene from Streptomyces scabiei 87.22; ASPFU_csn, chitosanase II from Aspergillus fumigatus Y2K; ASP_ORY_B, CsnB from Aspergillus oryzae IAM2660; HYPO_LIX, chitosanase from Trichoderma harzianum; META_ACR, Csn1 from Metarhizium acridum FI-985; FUS_SOL, chitosanase from Fusarium solani SUF386; NEURO, chitosanase from Neurospora crassa OR74A.

Directed mutagenesis experiments performed on the chitosanase from Fusarium solani f. sp. phaseoli SUF386 (110) as well as that of Aspergillus fumigatus (107) demonstrated that these chitosanases operate via an inverting mechanism and use a pair of carboxylic acid as catalytic residues.

The chitosanases from F. solani SUF386 and A. oryzae end with a cysteine-rich module which is not present in the other enzymes belonging to GH 75 family.

We can only speculate about the structural properties of family 75 enzymes. However, predictions of the secondary structure from Fusarium solani SUF386 chitosanase and those of Streptomyces avermitilis by an algorithm (111) reveal significant differences. These predictions suggest a high propensity for the formation of beta-sheets in the case of F. solani while the same algorithm predicts a rich helical structure for both S. avermitilis chitosanases.

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This page was created by Ryszard Brzezinski, Marie-Ève Lacombe-Harvey and Andrzej Neugebauer.
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Last updated: February 2014