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Glycoside Hydrolase Family 19

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Glycoside Hydrolase Family GH19
Clan none
(lysozyme fold)
Mechanism inverting
Active site residues known
CAZy DB link

Substrate specificities

Glycoside hydrolases of family 19 are endo-acting enzymes that hydrolyze glycosidic bonds within chitin, an insoluble polymer of β-1,4-linked N-acetyl-D-glucosamine (GlcNAc) and are thus referred to as chitinases (EC 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 β-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 CBM [2, 3]. Detailed studies on GH19 chitinases from Streptomyces (class IV; [3]) and rice (Oryza sativa; Class I; [4]) have 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 analysis[6]. Both structural characteristics (see below) and available biochemical data [3, 4] suggest that GH19 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

Conserved residues surrounding the catalytic acid, Glu67, in the family 19 papaya chitinase (PDB ID 3cql). Amino acid side chains are shown in stick representation. Hydrogen bonds are shown as dashed lines with distances indicated in Å. A GlcNAc2 unit binding in the -1 and +1 subsites is shown in narrow stick representation and an arrow indicates the position of the glycosidic oxygen. The sugar ring substituents are not shown in order to make the figure clearer. The structure of this complex comes from a model described in reference [9]. The picture is taken from reference [10].

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 general acid, whereas another glutamate located in a more variable loop-like structure (Glu89 in the barley enzyme) acts as the catalytic general base[9, 11, 12, 13].

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 [14] (see Figure). Interestingly, a similarly complex electrostatic interaction network is present in family GH46 chitosanases [15, 16] 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 GH46 chitosanases and with lysozymes in families GH22, GH23 and GH24 of glycoside hydrolases [17]. For a long time, structural information for these chitinases was limited to the structures of two class II plant enzymes [11, 18]. Recently, the structures of bacterial family GH19 chitinases have become available ([13, 19]; class IV), as well as the structures of class I [20] and class IV [21] GH19 chitinases from plants.

The structures of bacterial GH19 chitinases revealed several differences from the previously reported plant structures ([13, 19]; 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 [20, 22].

There is no structural information for GH19 enzymes in complex with their substrate. In 2008, Huet et al published [9] 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.

Structure images

Structures of the barley GH19 chitinase (left; PDB ID 1cns)and ChiG from S. coelicolor (right; PDB ID 2cjl). The side chains of the catalytic acids are shown in green. Additionally, the side chains of several residues that are (putatively) involved in substrate binding and catalysis are shown in red and purple. The picture is taken from reference [13].
Structural superposition of barley GH19 chitinase (light blue ribbon) and ChiG from S. coelicolor (darker blue surface). The picture highlights structural differences between the enzymes.The side chains of the catalytic glutamates in the bacterial enzymes are shown and labelled. The picture is taken from reference [13].

Family Firsts

First primary sequence determination
Bean leaf chitinase [23]
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 [12], structural analysis [11] and modelling [24]. Additional support from structure determination and modelling of a papaya chitinase [9].
First general acid residue identification
Chitinase from barley; determination by site-directed mutagenesis [12], structural analysis [11] and modelling [24]. Additional support from structure determination and modelling of a papaya chitinase [9].
First 3-D structure
Barley chitinase [11].


  1. Ohno T, Armand S, Hata T, Nikaidou N, Henrissat B, Mitsutomi M, and Watanabe T. (1996). A modular family 19 chitinase found in the prokaryotic organism Streptomyces griseus HUT 6037. J Bacteriol. 1996;178(17):5065-70. DOI:10.1128/jb.178.17.5065-5070.1996 | PubMed ID:8752320 [Ohno1996]
  2. Kawase T, Yokokawa S, Saito A, Fujii T, Nikaidou N, Miyashita K, and Watanabe T. (2006). Comparison of enzymatic and antifungal properties between family 18 and 19 chitinases from S. coelicolor A3(2). Biosci Biotechnol Biochem. 2006;70(4):988-98. DOI:10.1271/bbb.70.988 | PubMed ID:16636468 [Kawase2006]
  3. Heggset EB, Hoell IA, Kristoffersen M, Eijsink VG, and Vårum KM. (2009). Degradation of chitosans with chitinase G from Streptomyces coelicolor A3(2): production of chito-oligosaccharides and insight into subsite specificities. Biomacromolecules. 2009;10(4):892-9. DOI:10.1021/bm801418p | PubMed ID:19222164 [Heggset2009]
  4. Sasaki C, Vårum KM, Itoh Y, Tamoi M, and Fukamizo T. (2006). Rice chitinases: sugar recognition specificities of the individual subsites. Glycobiology. 2006;16(12):1242-50. DOI:10.1093/glycob/cwl043 | PubMed ID:16957091 [Sasaki2006]
  5. Fukamizo T, Koga D, Goto S Comparative biochemistry of chitinases - anomeric form of the reaction products Bioscience, Biotechnology & Biochemistry 1995, 59:311-313.

  6. Iseli B, Armand S, Boller T, Neuhaus JM, and Henrissat B. (1996). Plant chitinases use two different hydrolytic mechanisms. FEBS Lett. 1996;382(1-2):186-8. DOI:10.1016/0014-5793(96)00174-3 | PubMed ID:8612749 [Iseli1996]
  7. Honda Y and Fukamizo T. (1998). Substrate binding subsites of chitinase from barley seeds and lysozyme from goose egg white. Biochim Biophys Acta. 1998;1388(1):53-65. DOI:10.1016/s0167-4838(98)00153-8 | PubMed ID:9774706 [Honda1998]
  8. Sasaki C, Itoh Y, Takehara H, Kuhara S, and Fukamizo T. (2003). Family 19 chitinase from rice (Oryza sativa L.): substrate-binding subsites demonstrated by kinetic and molecular modeling studies. Plant Mol Biol. 2003;52(1):43-52. DOI:10.1023/a:1023972007681 | PubMed ID:12825688 [Sasaki2003]
  9. Huet J, Rucktooa P, Clantin B, Azarkan M, Looze Y, Villeret V, and Wintjens R. (2008). X-ray structure of papaya chitinase reveals the substrate binding mode of glycosyl hydrolase family 19 chitinases. Biochemistry. 2008;47(32):8283-91. DOI:10.1021/bi800655u | PubMed ID:18636748 [Huet2008]
  10. Hoell IA, Vaaje-Kolstad G, Eijsink VGH Structure and function of enzymes acting on chitin and chitosan Biotechnology and Genetic Engineering Reviews, Vol. 27, 2010, in press.

  11. Hart PJ, Monzingo AF, Ready MP, Ernst SR, and Robertus JD. (1993). Crystal structure of an endochitinase from Hordeum vulgare L. seeds. J Mol Biol. 1993;229(1):189-93. DOI:10.1006/jmbi.1993.1017 | PubMed ID:8421299 [Hart1993]
  12. Andersen MD, Jensen A, Robertus JD, Leah R, and Skriver K. (1997). Heterologous expression and characterization of wild-type and mutant forms of a 26 kDa endochitinase from barley (Hordeum vulgare L.). Biochem J. 1997;322 ( Pt 3)(Pt 3):815-22. DOI:10.1042/bj3220815 | PubMed ID:9148754 [Andersen1997]
  13. Hoell IA, Dalhus B, Heggset EB, Aspmo SI, and Eijsink VG. (2006). Crystal structure and enzymatic properties of a bacterial family 19 chitinase reveal differences from plant enzymes. FEBS J. 2006;273(21):4889-900. DOI:10.1111/j.1742-4658.2006.05487.x | PubMed ID:17010167 [Hoell2006]
  14. Ohnishi T, Juffer AH, Tamoi M, Skriver K, and Fukamizo T. (2005). 26 kDa endochitinase from barley seeds: an interaction of the ionizable side chains essential for catalysis. J Biochem. 2005;138(5):553-62. DOI:10.1093/jb/mvi154 | PubMed ID:16272567 [Ohnishi2005]
  15. Fukamizo T, Juffer AH, Vogel HJ, Honda Y, Tremblay H, Boucher I, Neugebauer WA, and Brzezinski R. (2000). Theoretical calculation of pKa reveals an important role of Arg205 in the activity and stability of Streptomyces sp. N174 chitosanase. J Biol Chem. 2000;275(33):25633-40. DOI:10.1074/jbc.M002574200 | PubMed ID:10829022 [Fukamizo2000]
  16. Lacombe-Harvey ME, Fukamizo T, Gagnon J, Ghinet MG, Dennhart N, Letzel T, and Brzezinski R. (2009). Accessory active site residues of Streptomyces sp. N174 chitosanase: variations on a common theme in the lysozyme superfamily. FEBS J. 2009;276(3):857-69. DOI:10.1111/j.1742-4658.2008.06830.x | PubMed ID:19143844 [Lacombe2009]
  17. Monzingo AF, Marcotte EM, Hart PJ, and Robertus JD. (1996). Chitinases, chitosanases, and lysozymes can be divided into procaryotic and eucaryotic families sharing a conserved core. Nat Struct Biol. 1996;3(2):133-40. DOI:10.1038/nsb0296-133 | PubMed ID:8564539 [Monzingo1996]
  18. Hahn M, Hennig M, Schlesier B, and Höhne W. (2000). Structure of jack bean chitinase. Acta Crystallogr D Biol Crystallogr. 2000;56(Pt 9):1096-9. DOI:10.1107/s090744490000857x | PubMed ID:10957628 [Hahn2000]
  19. Kezuka Y, Ohishi M, Itoh Y, Watanabe J, Mitsutomi M, Watanabe T, and Nonaka T. (2006). Structural studies of a two-domain chitinase from Streptomyces griseus HUT6037. J Mol Biol. 2006;358(2):472-84. DOI:10.1016/j.jmb.2006.02.013 | PubMed ID:16516924 [Kezuka2006]
  20. Ubhayasekera W, Tang CM, Ho SWT, Berglund G, Bergfors T, Chye ML, and Mowbray SL. (2007). Crystal structures of a family 19 chitinase from Brassica juncea show flexibility of binding cleft loops. FEBS J. 2007;274(14):3695-3703. DOI:10.1111/j.1742-4658.2007.05906.x | PubMed ID:17608716 [Ubhayasekera2007]
  21. Ubhayasekera W, Rawat R, Ho SW, Wiweger M, Von Arnold S, Chye ML, and Mowbray SL. (2009). The first crystal structures of a family 19 class IV chitinase: the enzyme from Norway spruce. Plant Mol Biol. 2009;71(3):277-89. DOI:10.1007/s11103-009-9523-9 | PubMed ID:19629717 [Ubhayasekera2009]
  22. Fukamizo T, Miyake R, Tamura A, Ohnuma T, Skriver K, Pursiainen NV, and Juffer AH. (2009). A flexible loop controlling the enzymatic activity and specificity in a glycosyl hydrolase family 19 endochitinase from barley seeds (Hordeum vulgare L.). Biochim Biophys Acta. 2009;1794(8):1159-67. DOI:10.1016/j.bbapap.2009.03.009 | PubMed ID:19332152 [Fukamizo2009]
  23. Broglie KE, Gaynor JJ, and Broglie RM. (1986). Ethylene-regulated gene expression: molecular cloning of the genes encoding an endochitinase from Phaseolus vulgaris. Proc Natl Acad Sci U S A. 1986;83(18):6820-4. DOI:10.1073/pnas.83.18.6820 | PubMed ID:2428042 [Broglie1986]
  24. Brameld KA and Goddard WA 3rd. (1998). The role of enzyme distortion in the single displacement mechanism of family 19 chitinases. Proc Natl Acad Sci U S A. 1998;95(8):4276-81. DOI:10.1073/pnas.95.8.4276 | PubMed ID:9539727 [Brameld1998]

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