Research Biochemistry and Glycobiology

Research: General information

Research within the laboratory of Biochemistry and Glycobiology involves a multidisciplinary study of carbohydrate-binding proteins (lectins) and some other plant proteins that are capable of interacting with endogenous or foreign (macro)molecules. The emphasis of the research is on protein-carbohydrate interaction and their involvement in signalling processes in plants or in plant protection. Based on the generated fundamental knowledge novel strategies are being developed to protect crop plants against pests and diseases. In addition, medical and therapeutic applications of lectins are investigated. Disciplines covered by the research projects include biochemistry and glycobiology as well as plant molecular biology, physiology, cell biology and genetics.

Plant lectins - a group of bioactive plant proteins

Many plants including important food plants such as wheat, potato, tomato and bean contain carbohydrate-binding proteins commonly referred to as "lectins", "agglutinins" or "hemagglutinins". This group of proteins comprises all plant proteins possessing at least one non-catalytic domain that binds reversibly to specific mono- or oligosaccharides. Hitherto, about 500 different plant lectins have been isolated and (partially) characterized. On a first glance, all these lectins form a heterogeneous group of proteins because of the obvious differences in structure, specificity and biological activities. Recently however, plant lectins have been classified on the basis of structural analyses and sequence data in twelve families of structurally and evolutionary related proteins. At present there are still a lot of unanswered questions regarding the physiological role of plant carbohydrate-binding proteins. Nevertheless, during the last decade some substantial progress has been made in our general understanding of the role of some plant lectins which are constitutively expressed in reasonable quantities.

Biochemical and molecular studies of numerous lectins demonstrated that only a limited number of carbohydrate-binding motifs evolved in plants. Since the specificity of these binding motifs is primarily directed against foreign glycans, it is generally accepted now that many plant lectins are involved in the recognition and binding of glycans from foreign organisms, and accordingly play a role in plant defense. Animal and insect feeding studies with purified lectins and experiments with transgenic plants confirmed that at least some lectins enhance the plants’ resistance against herbivorous higher animals or phytophagous invertebrates. To reconcile the presumed defensive role with the high concentration, the concept was developed that many plant lectins are storage proteins which, in case the plant is challenged by a predator, can be used as aspecific defense proteins. Evidently such a defense/storage role applies only to lectins that are present in relatively high concentrations.

In the last decade evidence accumulated that plants synthesize well-defined carbohydrate-binding proteins (lectins) upon exposure to stresses like drought, high salt, wounding, treatment with some plant hormones or pathogen attack. In contrast to the classical lectins that are often found in the plant vacuole this novel group of plant lectins locates to the cytoplasm and the nucleus of the plant cell. At present, a novel concept is being developed that these cytoplasmic/nuclear proteins have a specific endogenous role, and play an important role in normal growth and development of the plant through specific protein-carbohydrate interactions with regulatory glycoproteins in the cytoplasm and the nucleus.

Our current research focuses on the following topics:

  • Nictaba-related proteins
  • Protein degradation through protein-carbohydrate interactions
  • Proteins with an Euonymus lectin-like domain
  • Study of ribosome-inactivating proteins from edible fruits and vegetables
  • Molecular evolution of plant lectins
  • Use of lectins as resistance factors against pathogens and invertebrates

Nictaba-related proteins

Tobacco plants subjected to methyl jasmonate treatment accumulate an N-acetylglucosamine (GlcNAc) binding lectin called Nicotiana tabacum agglutinin, abbreviated as Nictaba. Nictaba is expressed in the cytoplasm and the nucleus of the parenchyma cells, where it interacts with glycosylated proteins. In search of the physiological role of Nictaba in the plant cell, a proteomics approach was used to purify and identify putative Nictaba-interacting proteins. We were able to show that Nictaba primarily associates with the core histone proteins. The co-localization and in vivo interaction between Nictaba and histone proteins was confirmed by microscopy based techniques. It is proposed that Nictaba might fulfill a signaling role in response to stress, by interacting with proteins in the plant cell nucleus.

Nictaba-like proteins have been identified in several plant species and are being investigated with respect to their biological activities and physiological role in the plant.

Protein degradation through protein-carbohydrate interactions

Accurate monitoring of protein quality control is of crucial importance for cell survival. A conserved proteolytic pathway involves the endoplasmic reticulum (ER)-associated degradation pathway, also known as the ERAD system. Herein, improperly folded (glyco)proteins or incompletely assembled oligomers are retained in the ER and translocated to the cytosol where they become labelled with a chain of ubiquitin molecules and subsequently recognized and degraded by the 26S proteasome. A crucial step in this pathway – the selective recognition of target proteins – is mediated by F-box proteins. Recently, a small group of mammalian F-box proteins (Fbs proteins) have been documented that share a sugar-binding domain which is involved in recognition and binding of glycoproteins that fail to achieve proper folding or assembly. After binding of these Fbs proteins with a misfolded glycoprotein, the glycoprotein is ubiquitinated and degraded by the proteasome. In this research project we are studying a new group of plant-specific Fbs-like proteins that are composed of an F-box domain linked to a Nictaba-like domain and that are assumed to be involved in (nuclear) glycoprotein degradation in plants.

Proteins with a Euonymus lectin-like domain

The molecular cloning of the Euonymus europaeus agglutinin allowed to classify this lectin into a new lectin family, further referred to as the group of proteins with an Euonymus lectin-like domain. Searches in the publicly available databases revealed that proteins with (an) EUL domain(s) are expressed in all Embryophyta. The family of EUL proteins is rather heterogeneous, in that some proteins consist of one EUL domain, while others comprise two in tandem arrayed EUL domains. Originally, these EUL proteins have been identified in rice, where they are expressed in the roots after treatment with abscisic acid and after exposure to salt-stress. The aim of our current research is to unravel the physiological importance of the EUL proteins in rice, Arabidopsis and Physcomitrella. Therefore the carbohydrate-binding specificity of the lectins is being analyzed and proteins interacting with EUL proteins are being investigated.

Study of ribosome-inactivating proteins from edible fruits and vegetables

Ribosome-inactivating proteins are a group of plant proteins which possess a highly specific N-glycosidase activity and are capable of catalytically inactivating eukaryotic ribosomes through the removal of a specific adenine residue from a highly conserved loop of the large rRNA. Ribosome-inactivating proteins (RIPs) from different plant species have received a lot of attention in biomedical research because they target conserved host protein synthesis machinery. RIPs offer a more or less pronounced protection in planta against viruses, fungi, and insects. In addition, many plant RIPs also show toxicity towards animal cells.

This project aims to study the biological activity and physiological role of RIPs from edible fruits, foods and vegetables.

Molecular evolution of plant lectins

Different carbohydrate-binding domains from plants are widespread within the plant kingdom. Using bioinformatics tools we have shown that these carbohydrate-binding modules have evolved during evolution. Next to lectins composed of carbohydrate-binding subunits only, a lot chimeric proteins have been identified where the carbohydrate-binding domain is linked to another unrelated domain.

Use of lectins as resistance factors against pathogens and invertebrates

Lectins have been attributed in plant defense against pathogens and phytophagous insects. The protective activity of lectins is in accordance with the observation that many plant lectins are not targeted against plant carbohydrates, but preferentially bind foreign glycans. Although evidence shows that the carbohydrate-binding activity of lectins is necessary for their insecticidal activity, the mode of action of lectins in insects remains enigmatic. Our research focuses on the insecticidal activity of several plant and fungal lectins, and the elucidation of their binding targets in the insect body/cells.





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