Difference between revisions of "Glycoside Hydrolase Family 19"

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Author: ^^^Vincent Eijsink^^^
Responsible Curator: ^^^Vincent Eijsink^^^

Glycoside Hydrolase Family GHnn
Clan	GH-x
Mechanism	inverting
Active site residues	known
CAZy DB link
http://www.cazy.org/fam/GH19.html

Substrate specificities

Glycoside hydrolases of family 19 hydrolyze glycoside bonds in chitin, an insoluble polymer of beta-1,4-linked N-acetyl-D-glucosamine (GlcNAc) and are thus referred to as chitinases (EC 3.2.1.14). These enzymes were originally identified in plants. In an older classification system for plant chitinases, comprising both GH18 and GH19 chitinases, family 19 enzymes comprise classes I, II and IV. In 1996, the first bacterial family 19 chitinase was described [1]. In addition to cleaving chitin, GH19 chitinases cleave soluble oligomers of beta-1,4-linked N-acetyl-D-glucosamine. For some plant enzymes lysozyme activity has been demonstrated. Currently available data suggest that GH19 enzymes are not particularly effective in degrading crystalline chitin (compared to certain members of the GH18 chitinase family), especially enzymes that lack CBMs. On the other hand GH19 enzymes are highly active on chitosans (= partially deacetylated chitin) with high degrees of acetylation, even if they lack a ChBD [2, 3]. Detailed studies on GH19 chitinases from Streptomyces (class IV; [3]) and rice (Oryza sativa; Class I; [4]) have both revealed that productive binding requires a GlcNAc to be bound in subsites -2 and +1, whereas deacetylated GlcNAc (GlcN) is tolerated in subsites -1 and +2.

Kinetics and Mechanism

Family 19 enzymes employ an inverting mechanism, as determined by NMR [5] and HPLC [6]. Both structural characteristics (see below) and available biochemical data [3, 4] suggest that GH 19 chitinases are non-processive endo-acting enzymes. Kinetic data for the conversion of polymeric and oligomeric substrates have been described in several studies. In some studies, kinetic data have been used to derive subsite binding affinties (e.g. [7, 8]).

Catalytic Residues

The catalytic residues are two glutamates. Although there still is limited structural information underpinning details of the inverting catalytic mechanism, there is considerable support for the notion that a glutamate located at the end of the third alpha helix (Glu 67 in the barley enzyme) acts as the catalytic acid, whereas another glutamate located in a more variable loop-like structure (Glu89 in the barley enzyme) acts as the catalytic base [9, 10, 11, 12].

It has been shown that at least two more conserved charged residues are crucial for catalysis. These residues, Glu203 and Arg215 in barley chitinase, form a triad together with the catalytic acid Glu67 [13] (see Figure). Interestingly, a similarly complex electrostatic interaction network is present in family 46 chitosanases [14, 15] with whom the family 19 enzymes share some overall structural similarity (see below).

Three-dimensional structures

The catalytic domains of family 19 chitinases have a lysozyme-like fold with rather shallow substrate-binding grooves that are not particularly rich in aromatic residues (see Figure). The catalytic domains of family 19 chitinases share a common fold with family 46 chitosanases and with lysozymes in families 22, 23 and 24 of glycoside hydrolases [16]. For a long time, structural information for these chitinases was limited to the structures of two class II plant enzymes [9, 17]. Recently, the structures of bacterial family 19 chitinases have become available ([11, 18]; class IV), as well as the structures of class I [19] and class IV [20] GH19 chitinases from plants.

The structures of bacterial GH19 chitinases revealed several differences from the previously reported plant structures ([11, 18]; see Figure). Compared to plant enzymes, the bacterial enzymes lack a C-terminal extension and three loops, some of which are thought to be flexible [19, 21].

There is no structural information for GH19 enzymes in complex with their substrate. In 2008, Huet et al published [12] the structure of a complex of papaya family 19 chitinase with GlcNAc units bound in the -2 and +1 subsites. This structure has been used to build a plausible model of a complex with (GlcNAc)4. This is the first structure (half experimental, half modeled) of an enzyme-substrate complex.

Family Firsts

First primary sequence determination: Bean leaf chitinase [22]

First stereochemistry determination: Yam chitinase, by NMR [5] and Bean chitinase, by HPLC [6].

First general base residue identification: Chitinase from barley; determination by site-directed mutagenesis [10], structural analysis [9] and modelling [23]. Additional support from structure determination and modelling of a papaya chitinase [12].

First general acid residue identification: Chitinase from barley; determination by site-directed mutagenesis [10], structural analysis [9] and modelling [23]. Additional support from structure determination and modelling of a papaya chitinase [12].

First 3-D structure: Barley chitinase [9].

References

All Medline abstracts: PubMed

@@ Line 47: / Line 47: @@
 == Three-dimensional structures ==
-The catalytic domains of family 19 chitinases have a lysozyme-like fold with rather shallow substrate-binding grooves that are not particularly rich in aromatic residues (see Figure). The catalytic domains of family 19 chitinases share a common fold with family 46 chitosanases and with lysozymes in families 22, 23 and 24 of glycoside hydrolases (Holm and Sander, 1994; Hart et al., 1995; Monzingo et al., 1996). For a long time, structural information for these chitinases was limited to the structures of two class II plant enzymes (Hart et al., 1993; Hahn et al., 2000). Recently, the structures of bacterial family 19 chitinases have become available (Hoell et al., 2006; Kezuka et al., 2006, class IV), as well as the structures of class I (Ubhayasekera et al. 2007) and class IV (Ubhayasekera et al. 2009) GH19 chitinases from plants.
+The catalytic domains of family 19 chitinases have a lysozyme-like fold with rather shallow substrate-binding grooves that are not particularly rich in aromatic residues (see Figure). The catalytic domains of family 19 chitinases share a common fold with family 46 chitosanases and with lysozymes in families 22, 23 and 24 of glycoside hydrolases <cite>Monzingo1996</cite>. For a long time, structural information for these chitinases was limited to the structures of two class II plant enzymes <cite>Hart1993 Hahn2000</cite>. Recently, the structures of bacterial family 19 chitinases have become available (<cite>Hoell2006 Kezuka2006</cite>; class IV), as well as the structures of class I <cite>Ubhayasekera2007</cite> and class IV <cite>Ubhayasekera2009</cite> GH19 chitinases from plants.
-The structures of bacterial GH19 chitinases revealed several differences from the previously reported plant structures (Hoell et al., 2006; Kezuka et al., 2006; see Figure). Compared to plant enzymes, the bacterial enzymes lack a C-terminal extension and three loops, some of which are thought to be flexible (Ubhayasekera et al., 2007; Fukamizo et al., 2009).
+The structures of bacterial GH19 chitinases revealed several differences from the previously reported plant structures (<cite>Hoell2006 Kezuka2006</cite>; see Figure). Compared to plant enzymes, the bacterial enzymes lack a C-terminal extension and three loops, some of which are thought to be flexible <cite>Ubhayasekera2007 Fukamizo2009</cite>.
-There is no structural information for GH19 enzymes in complex with their substrate. In 2008, Huet et al published  the structure of a complex of papaya family 19 chitinase with GlcNAc units bound in the -2 and +1 subsites. This structure has been used to build a plausible model of a complex with (GlcNAc)4. This is the first structure (half experimental, half modeled) of an enzyme-substrate complex.
+There is no structural information for GH19 enzymes in complex with their substrate. In 2008, Huet et al published <cite>Huet2008</cite> the structure of a complex of papaya family 19 chitinase with GlcNAc units bound in the -2 and +1 subsites. This structure has been used to build a plausible model of a complex with (GlcNAc)4. This is the first structure (half experimental, half modeled) of an enzyme-substrate complex.
@@ Line 90: / Line 90: @@
 #Fukamizo2000 pmid=10829022
 #Lacombe2009 pmid=19143844
-#
+#Monzingo1996 pmid=
-#
+#Hahn2000 pmid=
+#Kezuka2006 pmid=
+#Ubhayasekera2007 pmid=
+#Ubhayasekera2009 pmid=
+#Fukamizo2009 pmid=
+#Broglie1986 pmid=
+#Brameld1998 pmid=