Binding of the AVR4 Elicitor of Cladosporium fulvum to Chitotriose Units Is Facilitated by Positive Allosteric Protein-Protein Interactions

Abstract

The attack of fungal cell walls by plant chitinases is an important plant defense response to fungal infection. Anti-fungal activity of plant chitinases is largely restricted to chitinases that contain a noncatalytic, plantspecific chitin-binding domain (ChBD) (also called Hevein domain). Current data confirm that the race-specific elicitor AVR4 of the tomato pathogen Cladosporium fulvum can protect fungi against plant chitinases, which is based on the presence of a novel type of ChBD in AVR4 that was first identified in invertebrates. Although these two classes of ChBDs (Hevein and invertebrate) are sequentially unrelated, they share structural homology. Here, we show that the chitin-binding sites of these two classes of ChBDs have different topologies and characteristics. The K D , ⌬H, and ⌬S values obtained for the interaction between AVR4 and chito-oligomers are comparable with those obtained for Hevein. However, the binding site of AVR4 is larger than that of Hevein, i.e. AVR4 interacts strictly with chitotriose, whereas Hevein can also interact with the monomer N-acetylglucosamine. Moreover, binding of additional AVR4 molecules to chitin occurs through positive cooperative protein-protein interactions. By this mechanism AVR4 is likely to effectively shield chitin on the fungal cell wall, preventing the cell wall from being degraded by plant chitinases.Binding and conversion of carbohydrates by proteins is of fundamental importance in numerous biological processes, including (self and nonself) cell-cell recognition, cell adhesion, and carbohydrate turnover. Recently, protein domains responsible for this interaction have been reclassified into distinct carbohydrate-binding modules (CBMs) 1 (1). CBMs are often present in carbohydrate-degrading enzymes, where they appear to mediate a prolonged and more intimate contact between the catalytic domain and insoluble carbohydrate polymers (2, 3). Lectins, on the other hand, are carbohydratebinding proteins that lack enzymatic activity but often contain tandem repeats of CBMs.Chitin, a polymer consisting of ␤-1,4-linked GlcNAc residues, is a major component of crustacean shells, insect exoskeletons, and fungal cell walls but is absent in plants. In higher organisms, two CBMs predominantly confer binding of proteins to chitin, i.e. the Hevein domain (hereafter denoted as CBM18) (4) and the invertebrate chitin-binding domain (CBM14) (5). CBM18 is nearly exclusively found in plants (to date one additional member of CBM18 has been identified in Streptomyces griseus; Ref. 6), whereas CBM14 is commonly found in the genomes of baculoviridae, invertebrates, and mammals but absent in plants (5). Both ChBDs are typical CBMs, i.e. lectins with tandem repeats are known for both (e.g. wheat germ agglutinin (WGA) (7) and peritrophin-44 (8)), and both domains can be found in chitinases. However, CBM18 is only fused to the plant-specific family 19 catalytic domain, whereas chitinases of mammals and invertebrates utilize CBM14 in combination with the family 18 catalytic domain. Seque...

Tomato Cf resistance proteins mediate recognition of cognate homologous effectors from fungi pathogenic on dicots and monocots

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Genome sequence and virulence variation-related transcriptome profiles of Curvularia lunata, an important maize pathogenic fungus

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Identification of a Chitinase-modifying Protein from Fusarium verticillioides

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