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The leaf surface of most terrestrial plants is covered with plant hairs called trichomes. These epidermal appendages are thought to contribute to many aspects of plant defense against biotic and abiotic stresses in a variety of species. Trichome development has been intensively studied in Arabidopsis, and the phytochemical composition of trichomes was analyzed in a number of plant species. However, comparatively little is known of the proteins expressed. We therefore initiated a proteome approach to better define the cellular mechanisms operating in plant trichomes using two-dimensional gel electrophoresis to separate proteins of whole leaves and isolated trichomes. Tobacco was chosen due to the presence of glandular trichomes involved in the secretion of defense compounds. Comparative image analysis of the protein patterns indicated a number of spots, which were highly enriched in trichomes relative to leaves. These spots were excised for identification by mass spectrometry. The results showed that among the proteins specifically enriched in trichomes, the components of stress defense responses were strongly represented. The high expression of stress-related proteins was verified by Western blotting. Superoxide dismutase isoforms were additionally analyzed by activity staining. Our results demonstrate feasibility of the proteome approach to elucidate the cell biology of plant trichomes.

The structure of jack bean chitinase was solved at 1.8 A resolution by molecular replacement. It is an alpha-helical protein with three disulfide bridges. The active site is related in structure to animal and viral lysozymes. However, unlike in lysozyme, the architecture of the active site suggests a single-step cleavage. According to this mechanism, Glu68 is the proton donor and Glu90 assists in the reaction by moving towards the substrate and recruiting a water molecule that acts as the nucleophile. In this model, a water molecule was found in contact with Glu90 O(epsilon1) and Thr119 O(gamma) at a distance of 3.0 and 2.8 A, respectively. The model is in accordance with the observed inversion mechanism.

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The three-dimensional structure of narbonin, a seed protein from Vicia narbonensis L, has been determined at 1.8 A resolution. Phase information was obtained by multiple isomorphous replacement and optimized anomalous dispersion. The narbonin structure was initially traced with only 17% amino-acid sequence information and preliminarily refined to a crystallographic R-factor of 16.5%. It is now refined to 15.9% using full sequence information derived from cDNA and after the addition of more solvent molecules. The monomeric molecule of narbonin is an eight-stranded parallel beta-barrel surrounded by alpha-helices in a beta/alpha-topology similar to that first observed in triose phosphate isomerase. Differences exist in the N-terminal part of the polypeptide chain, where the first helix is replaced by a loop and the second beta-strand is followed by an additional antiparallel alpha-sheet placed parallel on top of alpha-helices alpha3 and alpha4. Two short additional secondary structures are present. The first, an alpha-helix, is situated between the seventh beta-strand and the following helix, and the second, which is a 3(10) helix, between the eighth strand and the C-terminal helix. The most striking observation is the lack of a known enzymatic function for narbonin, because all TIM-like structures known so far are enzymes.