Structural and dynamics studies of the protein elicitor cerato-platanin

Author(s)

Aline Lima de Oliveira

• Document type: Doctoral Thesis
• Press: Campinas, SP.
• Institution: Universidade Estadual de Campinas (UNICAMP). Instituto de Biologia
• Defense date: 2008-12-15

Examining board members

Thelma de Aguiar Pertinhez; Gonçalo Amarante Guimarães Pereira; Paula Regina Kuser Falcão; Marcius da Silva Almeida; Richard Charles Garratt

• Advisor: Celso Eduardo Benedetti; Thelma de Aguiar Pertinhez

Abstract

One of the most challenging problems facing plant pathology is to understand the molecular basis of the interaction between fungal plant pathogens and their host. Often, the mechanisms by which plants defend themselves against a variety of pathogens involve the recognition of specific molecules encoded by these microorganisms. Cerato-platanin (CP) is a moderately hydrophobic protein that is secreted and localized in the cell wall of Ceratocystis fimbriata produced by the ascomycete fungus Ceratocystis fimbriata, which is the causative agent of canker stain, a severe disease with incidence in a great number of plants, such as Platanus acerifolia, Theobroma cacao, Coffea Arabic, among others. The protein secreted by culture of Ceratocystis fimbriata f. sp. platani in the medium is able to elicit phytoalexin synthesis in Platanus acerifolia leaves and to enhance cell death. According to protein family database CP has been indicated as the founder of the "Cerato-Platanin Family" that includes other secreted fungal proteins involved in a variety of phytopathological phenomena and/or immunological reactions. The search in the Protein Data Bank revealed this protein family is not structurally characterized. In this way, the resolution of 3D structure of molecules involved in the host-fungus interaction is important to the comprehension of the initial events in the plant defence response. CP has been cloned and expressed in eukaryotic systems in its 13C, 15N isotopically labelled form. Multinuclear and multidimensional nuclear magnetic resonance has been used to determine its 3D solution structure. The CP structure is composed of two ß-strand parallel and five ß-strands antiparallel, assembled in a seven-stranded ß-sheet, facing three helical elements. The CP dynamics has been determined and the experimental data reveal that CP shows a structure characterized by limited internal mobility. Moreover, the protein has a good stability, keeping its conformation even at high temperature or in a wide rage of both pH and urea concentration. With the aim to search for other protein or domains conformationally similar to the CP, its 3D structure was submitted to structural analysis using ProFunc Server. The results of structure alignment show that CP does not share 3D structural similarity with any known avirulence proteins or elicitors from bacterial or fungal pathogens. However, surprisingly, the core of CP has a fold similar to molecules belonging to the chemokines and to the major histocompatibility complex class I (MHC-I) families. Indeed, the minimum energy CP structure superposes better to the dimeric form of interleunkin-8 (IL-8) and of the neutrophil activating peptide-2 (NAP-2), from the chemokines family, and to the a1, a2- domains of MHC-I protein. Interestingly, both families of proteins which have a similar fold to CP are related with defense response to pathogens in animals. The structural resemblance between CP and these defense signaling molecules suggests that, in the host cells, CP targets might have structural similarities to the human IL-8/NAP-2 or MHC-I proteins receptors and a search in the Arabidopsis genome with them produced a few hits. Not surprisingly, most of these proteins are surface or intracellular receptors implicated in the recognition of molecular patterns associated to pathogens or avirulence, a finding consistent with the hypothesis that CP, acting as an elicitor, should interact with such class of receptor proteins. In summary, although the biological function of the CP is still unknown, the elucidation of its 3D structure provides some insights about the molecular mechanism by which CP interact with their hosts. (AU)