Isolation, Partial Function Identification And Gene Cloning Of A Lectin From The Seeds Of Albizia Julibrissin

Plant lectins were defined as all plant protein possessing at least one non-catalytic domain, which binds reversibly to a specific mono- or oligosaccharide. Various functions of plant lectins were related with their sugar-recognition mechanisms. Since plant lectins were wide-spread through the plant kingdom and so many different lectins have been found, also their different biochemical properties, plant lectins would have much more applications, especially on pest control.This study reports the purification and some properties of a new lectin extracted from seeds of Mimosoideae, Albizia julibrissin, and also its antifungal activities. Based on this present studies, the seed lectin gene was cloned by RT-PCR and RACE from A. julibrissin and its structure and function was analyzed by bioinformatics. The main research conclusions were showed as follows:1. A new lectin was purified from Albizia julibrissin seeds by extraction, precipitation with (NH4)SO4, ion-exchange chromatography on DEAE Sepharose Fast Flow and followed native-PAGE. The purified lectin had a molecular mass of 25.4kDa on SDS-PAGE. Its isoelectric point is 7.96. The lectin was a glycoprotein . N-acetylglucosamine is the specific inhabitation sugar of the lectin.2. The function of this lectin showed that it strongly inhibited spore germination of Botrytis cinerea and Verticillium dahliae, and also partially inhibited the hyphal growth of Alternaria alternate. But it was no effect on the hyphal development of Fusarium oxysporum.3. A new lectin gene was cloned from Albizia julibrissin seed by RT-PCR and RACE. The full-length cDNA was 1096bp, and contained a 747bp open reading frame encoding a 248 amino acid protein with a 27 signal peptide.4. Protein structure was predicted with different methods. The results of primary structure analysis showed that the protein pI was 8.14, molecular weight was 26.8kDa. Using CLUSTAL (1.83) multiple sequence alignment methods determined Albizia julibrissin gene encoding amino acid sequence and related sequence homology. The amino acid sequence was found to be homologous to Mimosoideae, Parkia paltycephala, up to 85%.5. Related characters of coding protein were predicted. The result showed hydrophilic and transmembrane region located 12 to 34 amino acid residue. Secondary structure analysis indicated that the protein was mixed protein. Predicted three-dimensional structure of the protein by homology method in different net server and model quality was evaluated by PROCHECK software. The MODEL EsyPreD was BEST.6. Function analysis of the gene deduced amino acid sequence showed it belonged to Glyco-hydro₁8 superfamily. It had a domain of Glyco-hydro₁8 superfamily and located from 146 to 154 amino acid residue. The sequence was LDGVDFDIE. The subcellular location indicated the protein was extracellular protein, and had a transmembrane segment, the sequence was LVLQVCLIMMVSTAKAGGIVVYW.This data are great importance considering the lack of information on Mimosoideae plants. And also the conclusions provide some information for pest resistance research further in forest trees.