Molecular characterization of an enzymatic D-galactose specific lectin from field bean (Dolichos lablab) seeds.

Devavratha, H. Rao (2013) Molecular characterization of an enzymatic D-galactose specific lectin from field bean (Dolichos lablab) seeds. Doctoral thesis, University of Mysore.

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Abstract

The phenomenon of one “protein many functions” an emerging new concept is finding experimental support. Multifunctional proteins are proteins that have multiple catalytic or binding sites. The synonymous terms “multiheaded”, “polycephalic” or “chimeric” proteins imply that the active center and binding site are generated by the folding of contiguous stretches of the polypeptide chain to yield autonomous globular structures or domains. It is known that functional sites are located either between domains or at the inter-subunit interfaces. Thus, a protein active as an enzyme can be recruited to do other functions. These proteins may have multiple catalytic sites, binding properties and structural roles. The specific binding of different ligands to protein may also serve as regulatory signals. Preliminary evidences from this laboratory indicate that the Polyphenol oxidase (1, 2 benzene:oxygen oxidoreductase; EC 1.10.3.1, PPO) isolated from field bean (Dolichos lablab) is a multifunctional protein that displayed associated haemagglutinating activity. The protein exhibits catalytic activity by oxidizing diphenols to o-quinones and binding properties similar to that of Gal binding lectins. This protein was designated as PPO-haemagglutinin. Both PPO and lectin are plant pathogen related proteins (PRP), which are involved in plant defense against pathogen attack. PPO is a bifunctional copper containing enzyme, which in the presence of molecular oxygen catalyses the hydroxylation of monophenols to o-diphenols (monophenolase, cresolase activity) and further oxidizes the o-diphenols to o-quinones (diphenolase, catecholase activity). The generated, unstable highly reactive o-quinones subsequently react with themselves, amino acids or proteins evolving into brown, black or red heterogeneous polymers often referred to as melanins. The PPO mediated browning of fruits and vegetables’ following damage to the organism suggests a role in plant defense against insects and pathogens and is considered as a defense related antinutritive antioxidant enzyme. 1 Lectins are multivalent cell agglutinating proteins, which by virtue of their exquisite sugar binding specificities have wide spread applications. The legume lectins are a family of sugar binding proteins found in the seeds, and to a lesser extent in the stems and the leaves, of plants belonging to the Leguminosae family. As defense proteins they bind to the brush border membrane of the insect intestine or the glycoproteins in their peritrophic matrix laminar epithelial cells in the midgut and disrupt digestive processes and nutrient assimilation. New directions in plant lectin research beyond carbohydrate binding have viewed lectins as potential candidates for targeted drug delivery due to their ability to bind non-carbohydrate ligands. Compelling evidence as detailed by the sequence identity and a recent report of a Gal specific lectin with an NH2-terminal sequence and subunit architecture identical to the purified PPO led to the conclusion that field bean PPO is a Gal specific lectin, designated as PPO-haemagglutinin. The findings 1) the Gal specific lectin of field bean (Dolichos lablab) potent polyphenol oxidase activity and 2) the Gal specific lectin of horse gram (Dolichos biflorus) exhibits lipoxygenase activity leads to the question “are all Gal specific lectins enzymes”. Both the enzyme activities are related to plant defense; therefore how important are these proteins to the physiology of plants? Understanding the structure-function relationships of such proteins would not only provide valuable information on defense mechanism adaptations in plants, but also insights in to the mechanistics of multifunctionalities. Preliminary information on the functionality of the PPO-haemagglutinin from the seeds of Dolichos lablab indicates that the PPO and haemagglutination activities are associated with separate loci on the same protein. To complement and extend these studies to further understand how the protein acts as an oxidative enzyme and can also be recruited to act as a binding protein, calls for a detailed characterization at the molecular level. The outcome of such a study would be useful and lend support in elucidating the structural features that delineate the two functionalities 1) oxidase activity and 2) sugar binding properties.

Item Type:	Thesis (Doctoral)
Uncontrolled Keywords:	field bean, Dolichos lablab, Lectins, oxidative enzyme
Subjects:	500 Natural Sciences and Mathematics > 04 Chemistry and Allied Sciences > 16 Enzyme Chemistry 600 Technology > 08 Food technology > 22 Legumes-Pulses
Divisions:	Protein Chemistry and Technology
Depositing User:	Food Sci. & Technol. Information Services
Date Deposited:	17 Mar 2015 11:39
Last Modified:	25 Sep 2018 06:56
URI:	http://ir.cftri.res.in/id/eprint/11754

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